To investigate the underlying mechanisms of the observed temporal tuning properties of LSO and their sources of variability, we used a simple coincidence counting model and examined how specific parameters of coincidence detection affect monaural and binaural AM coding. Spike rates and phase-locking of evoked excitatory and spontaneous inhibitory inputs had only minor effects on LSO output to monaural AM inputs. In contrast, the coincidence threshold of the model neuron affected both the overall spike rates and the half-peak positions of the AM-tuning curve, whereas the width of the coincidence window merely influenced the output spike rates. The duration of the refractory period affected only the low-frequency portion of the monaural AM-tuning curve. Unlike monaural AM coding, temporal factors, such as the coincidence window and the effective duration of inhibition, played a major role in determining the trough positions of simulated binaural phase-response curves. In addition, empirically-observed level-dependence of binaural phase-coding was reproduced in the framework of our minimalistic coincidence counting model. These modeling results suggest that coincidence detection of excitatory and inhibitory synaptic inputs is essential for LSO neurons to encode both monaural and binaural AM sounds.Many sensory neurons encode temporal information by detecting coincident arrivals of synaptic inputs. In the mammalian auditory brainstem, binaural neurons of the medial superior olive (MSO) are known to act as coincidence detectors, whereas in the lateral superior olive (LSO) roles of coincidence detection have remained unclear. LSO neurons receive excitatory and inhibitory inputs driven by ipsilateral and contralateral acoustic stimuli, respectively, and vary their output spike rates according to interaural level differences. In addition, LSO neurons are also sensitive to binaural phase differences of low-frequency tones and envelopes of amplitude-modulated (AM) sounds. Previous physiological recordings  Detecting coincident arrivals of synaptic inputs is a shared fundamental property of many sensory neurons. Such 'coincidence detection' usually refers to the detection of synchronized excitatory inputs only. Experimental evidence, however, indicated that some auditory neurons are also sensitive to the relative timing of excitatory and inhibitory synaptic inputs. This type of sensitivity is suggested to be important for coding temporal information of amplitude-modulated sounds, such as speech and other naturalistic sounds. In this study, we used a minimal model of coincidence detection to identify the key elements for temporal information processing. Our series of simulations demonstrated that (1) the threshold and time window for coincidence detection were the major factors for determining the response properties to excitatory inputs, and that (2) timed interactions between excitatory and inhibitory synaptic inputs are responsible for determining the temporal tuning properties of the neuron. These results suggest that coincidence detection is an essential function of neurons that detect the 'anti-coincidence' of excitatory and inhibitory inputs to encode temporal information of sounds. Sound source localization, or the ability to determine the direction of a sound source, is a fundamental function of the auditory system. The interaural level difference (ILD), which is the difference of the sound levels between the two ears, is one of the primary cues of azimuthal sound localization . ILDs gAmplitude modulation is a general feature of natural stimuli including communication sounds, and hence processing of AM signals is a fundamental property of the auditory system . For exafm) of 100–500 Hz, and gradually decreased down to 50 spikes/sec at around fm = 600–1000 Hz  conductance led to rate-fm functions that were more consistent with empirical results. Nevertheless, no combinations of parameters were able to comprehensively explain the diversity of LSO responses to monaural AM stimuli.LSO responses to monaural AM tones at different modulation frequencies were systematically studied by Joris and Yin . Typicaljections ,38. Wangjections  studied jections , was unlin vivo revealed a number of characteristic response properties of LSO neurons to binaural AM sounds [fm , wherIn order to reveal underlying mechanisms of monaural and binaural AM coding in the LSO, we use a simple model of coincidence detection, which is based on the counting of coincident inputs ,43. Becat is time, p is a 2π-periodic function, and fm is the stimulus modulation frequency. We used a von-Mises distribution function [p(x). Assuming that In is the Modified Bessel function of order n, the periodic function we used is written as:k being the concentration parameter that defines the steepness of the distribution. The degree of phase-locking, measured as vector strength (VS) [Bushy cells in the VCN generally show phase-locked spiking activities to the envelope of AM sound stimuli. Excitatory phase-locked synaptic inputs to the LSO were modeled as an inhomogeneous Poisson process ,47. The function  for the gth (VS) , is relaFor more detail about theoretical formulations, see .fm being the modulation frequency in Hz. Since there were considerable variations in bushy cell responses between animal species and between experimental conditions, we used a simple linear function for the rate (0 (input intensity at zero modulation frequency) was 180 (spikes/sec). The non-linear decreasing function for simulating VS reaches zero at fm = 2000 Hz. In our simulations, we used the above-defined rate and VS functions unless otherwise stated. As in a previous study [The average spike rate and the VS of a bushy cell to AM stimuli generally decrease with the modulation frequency . Basthe rate  and a mothe rate , both of1−exp(fm−00/500))/us study , we assuλinh = 30 (spikes/sec). We assumed that an LSO neuron receives 8 inhibitory inputs [Even without sound stimuli, LSO neurons receive spontaneous inhibition from MNTB neurons ,50. Basey inputs . For test, the coincidence threshold between time t and t+Δ was elevated by δ. We refer to this effective length Δ of the inhibition as 'inhibition window' and to the elevation δ of the threshold as 'threshold increase'. Standard parameter values are summarized in Coincidence detection in the LSO was modeled as simple coincidence counting , which wOur coincidence counting model has three fundamental parameters: the coincidence threshold θ, the width of the coincidence window W, and the length of the refractory period T. The default values we used were: θ = 8 inputs, W = 0.8 ms, and T = 1.6 ms, which were taken from empirically measured parameter ranges  to reproTo test combined effects of the coincidence threshold and window, we varied these parameters together while fixing the ratio W/θ, so that the average number of required inputs within the window remains unchanged. We also covaried these parameters so that the maximum response rate was constant , where R is the VS of the LSO output spike sequence at the modulation frequency fm. Following a notation of prior studies , we alsfm = 300 Hz, which roughly corresponded to the peak of the monaural AM response  and decreasing (at high levels) parts of the curve, respectively  and Ni(t) be the numbers of excitatory and inhibitory inputs, respectively, that the model neuron received between the end of the last refractory period and time t. When the summed excitatory spike counts subtracted by the weighted number of inhibitory inputs reached or exceeded the pre-set threshold θ, one output spike was generated and the model was then in a refractory period T. Namely, a spike was generated at time t, when Ne(t) - δNi(t) ≥ θ. The weight δ of the inhibitory input corresponds to the threshold increase by inhibition. Thus the pure integrator model has three parameters: threshold θ, refractory period T and threshold increase δ. We used the same default values for this model  as for the coincidence counting model  modulation frequencies.Next, effects of the refractory period T were tested . SimulatOur simulation results showed that spontaneous inhibitory inputs had only limited effects on monaural AM tuning . In binaThe effect of the threshold increase δ was most prominent when it was varied from zero  to one or two, resulting in a transition from a flat to standard phase-response curve with a distinct peak and trough . FurtherVariation in the width Δ of the inhibition window showed more dynamic effects on phase-response curves . A narrofm. A high modulation frequency generally led to lower spiking rates, which was shown in previous in vivo recordings .In the preceding sections, we investigated monaural and binaural AM coding in the LSO with fixed sound stimulus levels. Since LSO neurons are sensitive to binaural level differences, we here examine how ILDs affect the output of our coincidence counting model. We varied ILDs of non-modulating inputs . Simulatin vivo recording data from the cat LSO , suggesOur simulation results suggest that temporal processing of excitatory and inhibitory inputs is essential for both monaural and binaural AM-tunings. To further confirm the importance of coincidence detection in LSO, we performed additional simulations using a pure integrator model fed with the same set of inputs as used for the coincidence counting model. The perfect integrator model does not have a coincidence window or an inhibition window, since it simply sums excitatory inputs and subtracts inhibitory inputs regardless of their timings see .Simulated rate-MTF curves of the pure integrator showed a broad tuning , which rFor binaural inputs, the pure integrator model also showed sensitivity to time differences of excitatory and inhibitory inputs . Althougin vivo [Amplitude modulation is a common feature of natural stimuli, and hence processing of AM signals is important in auditory  and othein vivo . Furtherin vivo .Octopus cells in the posteroventral cochlear nucleus and principal cells of the MSO are also known to act as auditory coincidence detectors . The intMonaural coincidence detection has been studied frequently in relation to the improvement of temporal coding performance at the cochlear nucleus ,66,67, win vivo needs to be tested in future experiments.The parameters of excitatory inputs , which depended on the modulation-frequency, had only limited effects on monaural AM coding . SpontanIn contrast to monaural AM-tuning curves, binaural phase-response curves showed distinct dependence on the model parameters. Both the coincidence threshold and window affected the LSO output rates to simulated binaural inputs, but the trough position of a phase-response curve was not sensitive to the threshold but to the window width . Moreovein vivo, for example, ranged from 1.1–2.8 ms [in vitro showed variations in the refractory period [in vitro also showed considerable variations (9.6 ± 2.8 excitatory inputs)[In agreement with our results, previous experimental studies reported comparable variations in coincidence detection parameters across neurons. The refractory period of cat LSO neurons 1–2.8 ms . Similary period . As prevy period . The numy inputs). The widy inputs), such asy inputs), inhibity inputs)) and deny inputs). Excitaty inputs), which sy inputs); 1.9–4.1y inputs); and 8.5y inputs)). A prevy inputs).in vitro, for example, might reveal more detailed relations between the higher-level coincidence parameters and the underlying membrane and synaptic factors. Previous studies showed that having non-homogeneous neuronal population is often beneficial for coding sensory information [The KLVA conductance, a major determinant of the resting membrane properties , shows ormation ,80,81. TOur minimalistic coincidence counting model has only a small number of parameters , which mIn our phase-tuning curves Figs –12, modeA prior modeling study suggested that a large amount of KLVA conductance was necessary to reproduce empirical AM-frequency dependence of LSO spike rates . The majThe coincidence counting model we used was originally introduced for investigating the binaural coding in the MSO . This mofm rates [in vivo methods, which were recently used for MSO neurons [Although our results were consistent with previous empirical data ,28,36, mfm rates , but onlfm rates ,98. Timefm rates . How inh neurons .in vivo showed that coincidence detection in MSO is affected by its input history [Our LSO model consisted of two parts: (1) generation of time-locked excitatory and inhibitory inputs and (2) coincidence detection of the LSO neuron. As was done in a previous modeling study , sound-d history . Similar


We assessed whether the reported decrease in fertility rates among 15 to 19 years old Brazilian adolescents has met with a parallel decrease in very young adolescent (10 to 14 years old) fertility rates. So we explored temporal trends for fertility rates among very young adolescents between 2000 and 2012 for Brazil as a whole, its regions and states; and also analyzed the spatial distribution of fertility rates among Brazilian municipalities in the years 2000 and 2012.We used data from the Information System on Live Births to calculate the rates. To examine the temporal trends, we used linear regression for time series with Prais-Winsten estimation, including the annual percentage change, for the country, regions, and states. To analyze the spatial distribution among Brazilian municipalities, we calculated the Global Moran Index and created a local Moran significance and cluster map through Local Indicators of Spatial Association (LISA). We also elaborated a thematic map with the rates using empirical Bayesian estimation.p = 0.05). The maps indicated a heterogeneous distribution of the rates, with high-rate clusters predominant in the North and low-rate clusters predominant in the South, Southeast, and Midwest.Brazilian very young adolescent fertility rates remained high and stable throughout the 2000 to 2012 period, and significantly decreased in three out of 26 states, and in the federal district. On the other hand, an increase was observed in two Northern and Northeastern states. The rates were spatially dependent in Brazilian municipalities (Moran Index = 0.22 in 2012; Our findings indicate that Brazilian very young adolescent fertility rates have not decreased in parallel with adolescent fertility rates as they remain high and did not decrease from 2000 and 2012, even though a few states presented a decrease. Thus, these phenomena probably have distinct underlying causes that warrant further elucidation. Progress in this field is crucial for the development of specific policies and programs focused on very young adolescents. Adolescent pregnancy has strong negative effects on the educational and professional opportunities for women . Thus, cWhere measured, childbirths among very young adolescents (VYA) aged 10 to 14 years occur at much lower rates. For example, while the adolescent fertility rate is 10 in Spain and 34 in the USA, the VYA fertility rates in these countries are only 0.14 and 0.45, respectively .In face of these drastic consequences and the lack of data, the World Health Organization  has emphBrazil is a federative republic that covers almost half of South America Fig. , with a The country has presented one of the highest levels of income inequality in the world . Even thWe analyzed the live births registered in Brazil from 2000 to 2012. In this period, there were 359,963 live births to women aged 10 to 14 years old, representing 0.93% of all 38,815,012 births at the national level [http://www2.datasus.gov.br/DATASUS/index.php).To estimate the VYA fertility rate, we divided the births to women aged 10 to 14 years by the number of female residents aged 10 to 14 years old. The values are presented in terms of live births per 1,000 adolescents. The numerators of the age-specific fertility rate for the VYA were provided by the Brazilian Information System on Live Births (SINASC), and the denominators were obtained from the National Bureau of Statistics (IBGE). For 2000 and 2010, we used censuses’ data. For the other years, we used projections. Regarding 2011 and 2012, the Ministry of Health/DATASUS applied the age/sex distribution observed in 2010 census to adjust data, since IBGE projected only total population by municipalities , 16. DatThe study protocol conforms to the ethical guidelines of the 1975 Declaration of Helsinki and was approved by the University of São Paulo School of Nursing, Brazil (no. 170.604).In order to chart the temporal trend between 2000 and 2012, we calculated the VYA fertility rates by running a linear regression for each of the 26 states, the federal district, the five regions  and the whole country. Trend analysis was performed in Stata 13.0 using linear regression for time series with Prais-Winsten auto-regression procedure to minimize first-order temporal autocorrelation of residues. The dependent variable was 10-to-14 fertility rates  and the independent variable was the calendar year (2000 to 2012). When the regression coefficient was not significantly different from zero (p > 0.05), the trend was considered stable. When the coefficient was positive or negative, the trend was considered as increasing and decreasing, respectively. We also calculated the annual percent change (APC) and its respective 95% confidence interval (CI).To analyze the spatial distribution of the fertility rates in the total of municipalities in the years 2000 and 2012, we calculated the rates for each of the 5,565 municipalities of Brazil. In 2000, there were six births with missing information about the birthplace, whereas in 2010 there was only one. These cases were excluded. We used digital maps from 1997 and 2013 available at the National Bureau of Statistics/IBGE . The 199p < 0.05). It is important to point out that both Global Moran Index and LISA analysis were conducted with ‘pure’ VYA fertility rates in order not to influence the estimates, even considering instability due to the great variability on the number of local live births and female 10–14 years-old population.We conducted spatial analysis in three ways. First, we used Global Moran Index to verify the presence of global spatial dependence on the age-specific fertility rates . Second,Third, we elaborated maps with local VYA fertility rates estimated with the Empirical Bayesian method, with the purpose of minimizing the bias of local level age-specific rate estimation as large variations are expected among very small municipalities –21. ThisFigure Table p = 0.05) for 2000 and 2012, respectively. These results show that fertility rates among the VYA are spatially dependent. In 2000, the LISA map  when compared to high-income countries such as Switzerland, Sweden, and New Zealand, where rates vary from 0.01 to 0.17 . HoweverSexual violence or coercion towards VYA may also play a role in determining the high fertility rate of this group in Brazil, as elsewhere. Intercourse in the VYA age group is more likely to be nonconsensual , and theSome significant decreases in VYA fertility rates were observed in regions and states with higher HDI including the South and Midwest regions, the states of Mato Grosso, Rio de Janeiro, and Rio Grande do Sul, and the federal district. Because the HDI integrates income, health and education indicators , it reflThe Midwest region displayed a significant decrease in fertility rates. However, one of the states of this region, where the rates remained stable – Mato Grosso do Sul – showed cluster areas formed in border regions with Paraguay and Bolivia. Border areas usually differ from other locations, because nationalities, cultures, and customs mingle. Moreover, border regions favor drug trafficking and the sexual exploitation of children and adolescents. This context may contribute to the occurrence of VYA pregnancies, as it does regarding the spread of HIV/AIDS , 35.Despite these scattered decreases in fertility, the high VYA fertility rates detected in the country and the increases observed in two states (Amazonas and Bahia) represent an important social concern. Amazonas is the largest Brazilian state, and most of its area is covered by the Amazon Forest. More than 178 thousand indigenous people live in protected reservations or unprotected areas . The strSexual tourism may explain the increase in VYA fertility rate observed in the state of Bahia. The map shows an increase in high-rate clusters in the south of the state, coinciding with the location of touristic municipalities. A study conducted at touristic seaside municipalities in the states of São Paulo and Rio de Janeiro indicated that tourism has some negative impacts on host communities, which become more susceptible to risky sexual behaviors and elevated alcohol use . The samThe states of Amazonas and Bahia also underperform in terms of investments in VYA education. The proportion of 15- to 17-year-old adolescents who finished middle school is less than 45%, similarly to other Northern and Northeastern states . DelayedThe analysis of the spatial distribution of fertility rates among Brazilian municipalities reiterates the pattern, with a concentration of higher rates in the Northern region, including the state of Amazonas, and some areas in the Northeast. These findings indicate that births among VYA may be associated with income inequality much as observed for older adolescents . DespiteSome limitations of our study should be pointed out. The rate denominators (number of girls aged 10–14) are intercensal estimates based on trends analysis (except for the years 2000 and 2010) and could influence the results. The use of secondary data from the Brazilian Information System on Live Births/SINASC may have inconsistencies, especially because reliability and underreporting may differ significantly among regions. Anyway, SINASC currently covers more than 90% of births in the whole country and shows substantial improvement in the last decade, especially in Northern and Northeastern regions , 38. RegOur results provide an ecological perspective. Additional research should address Brazilian VYA to generate demographic data on sexual and reproductive health such as age of first intercourse, contraceptive use, pregnancy, delivery and maternal morbidity and mortality, including ethnicity issues. Moreover, further research should be interdisciplinary, also covering cultural values, educational and professional perspectives, family environment, and connectedness to school.Our findings indicate that Brazilian VYA fertility rates have not decreased in parallel with adolescent fertility rates. Thus, these phenomena probably have distinct underlying causes that warrant further elucidation. Progress in this field is crucial for the development of specific policies and programs focused on VYA.The study protocol conforms to the ethical guidelines of the 1975 Declaration of Helsinki and was approved by the University of São Paulo School of Nursing, Brazil (no. 170.604).http://www2.datasus.gov.br/DATASUS/index.php) and IBGE (http://www.ibge.gov.br/home/).Data supporting our findings are of open access and can be found at DATASUS (


Clostridia, Enterobacteriaceae, and low G+C Gram-positive cocci. UA-degrading Clostridia had never been isolated from termite guts. UA-degrading ability was sporadically distributed among phylogenetically various culturable anaerobic bacteria from termite guts. A strain of Clostridium sp., which was commonly isolated from three termite species and represented a probable new species in cluster XIVa of clostridia, utilized UA as a nitrogen source but not as a sole carbon and energy source. This feature is in clear contrast to that of well-studied purinolytic clostridia or previously isolated UA degraders from termite guts, which also utilize UA as a sole carbon and energy source. Ammonia is the major nitrogenous product of UA degradation. Various purines stimulated the growth of this strain when added to an otherwise growth-limiting, nitrogen poor medium. The bacterial species involved the recycling of UA nitrogen in the gut microbial community of termites are more diverse in terms of both taxonomy and nutritional physiology than previously recognized.Recycling of the nitrogenous waste uric acid (UA) of wood-feeding termites by their gut bacteria is one of the significant aspects of symbiosis for the conservation of nitrogen sources. Diverse anaerobic UA-degrading bacteria comprising 16 species were isolated from the gut of eight termite species, and were assigned to  Not merely regarded as nitrogenous waste, UA is apparently utilized as a nitrogen source or metabolic reserve in some insects, particularly those existing on a nitrogen-poor diet (e.g.Uric acid (UA) is a major nitrogenous waste excreted from animals such as terrestrial insects, birds, and certain reptiles , Sebaldella (formerly Bacteroides) termitidis, and Citrobacter sp. Despite the repeated extensive isolation, however, only a single species of the subterranean termite Reticulitermes flavipes has been investigated. There are more than 270 termite genera and a marked difference in gut microbiota has been disclosed among genera (Clostridium (designated strain NkU-1) was further investigated for its properties of UA utilization.Uricolytic bacteria have been isolated from the termite gut, and well studied physiologically , 42. Theg genera , 56; the−4% containing 1.0 and 0.1% of UA and 0.6 and 1.5% agar in the top and bottom layers, respectively. The inoculated plate was anaerobically incubated at 25°C for 14 days. The colonies surrounded by a clear zone in the otherwise opaque medium were counted as presumptive uricolytic bacteria. Anaerobic manipulation and cultivation were performed in an anaerobic chamber  with an O2-free atmosphere of N2/H2/CO2 . In the case of the termite Reticulitermes speratus, we applied a different method to identify UA-degrading bacteria; anaerobic bacteria were first isolated in medium that did not contain UA, as described previously , with an 80% confidence threshold. For detailed comparisons, the quality-checked sequences of type strains in “The All-Species Living Tree” project (http://www.arb-silva.de/) and used. Phylogenetic analysis with the maximum likelihood method was performed with RAxML 7.2.6 , 0.1% yeast extract, 0.1% UA, 2.5% (v/v) 40× salt solution, and 0.4% (v/v) vitamin solution was also used for the culture of several strains to examine UA degradation. In the case of strain Cd23, GY instead of TYU medium was used for this purpose. UA was solubilized as 2% solution in 0.5 N NaOH, filter sterilized and added at an appropriate final concentraion, and before inoculation, a predetermined amount of sterile 0.5 N HCl was added to neutralize the medium. Purines to be added to the medium were solubilized as described above for UA and in the previous report , 2.5% (v/v) 40× salt solution, 5% (v/v) 20× KP buffer, 0.4% (v/v) vitamin solution, and 10s report . UA, pur3 was determined enzymatically by NH3-dependent oxidation of NADPH (measured by absorbance at 340 nm) with beef liver glutamate dehydrogenase (Roche Applied Science). UA and NH3 in growth media were assayed after removal of cells by centrifugation. All measurements were performed in duplicate and mean values are shown.In liquid culture, cell growth was measured by optical density (OD) at 660 nm or by the protein amount using Bio-Rad Protein Assay Reagent after adding NaOH  and boiling the culture. UA was measured spectrophotometrically by the difference in its specific absorbance at 292 nm before and after treatment with pig liver uricase (Toyobo). NH3 to 4.0×104 cells per gut except in the case of Neotermes koshunensis (1.1×106 cells per gut). Because of the low culturability of gut bacteria  was also examined, but uricolytic bacteria were not obtained, probably because its food (soil organic matter) is rich in nitrogenous compounds and recycling of UA is less significant in this termite.Anaerobic uricolytic bacteria were successfully detected on the BHIU plate from eight termite species at rates ranging 0.1 to 48.6% CFU . They cobacteria , 50), isClostridia, Enterobacteriaceae (Gammaproteobacteria), and low G+C Gram-positive cocci (Bacilli). To our knowledge, this is the first report for the isolation of uricolytic bacteria in Clostridia from termite guts, although a variety of Clostridia strains have been isolated  could not be isolated in this study.The uricolytic isolates were taxonomically classified, and 16 different species were identified . They weClostridia occurred in six of the eight examined termite species , which occurred commonly in two termites and frequently among uricolytic isolates from Nasutitermes takasagoensis, were closely related to C. sphenoides and C. celerecrescens (each 99.9% identity). Indeed, type strains of these two Clostridium species degraded UA in the BHIU plate medium as much as strains NkU-1 and Cd6 (determined by clear zone formation), whereas those of C. xylanolyticum and C. aerotorelance did not (data not shown). All of the isolates in Clostridia were distantly related to well-studied UA-degrading Clostridium species, C. acidiurici, C. purinolyticum (both belonging to cluster XII), and C. cylindrosporum (forms a unique lineage near cluster I or II)  , 49, 57.lostridia, althougR. speratus, various anaerobic bacteria affiliated to the genera Enterobacter, Clostridium, Lactococcus, Bacillus, and Bacteroides were isolated in addition to previously reported Disgonomonas sp. (Enterobacter sp. (strain RsN-1) could degrade UA on the BHIU plate. The isolates in Clostridium and Lactococcus from R. speratus corresponded to completely different species from the uricolytic strains from other termites. Furthermore, Lactococcus spp. are those of major culturable bacteria in termite guts, but only a few among numerous isolates have demonstrated UA-degrading ability (i.e. Lactococcus or Clostridium).From onas sp. . Neverth ability , 41, 50;Clostridium sp. strain NkU-1, its closely related three strains from the same termite, and Lactococcus sp. strain Cd31 degraded UA almost completely, whereas Enterococcus sp. strain Cd23, Enterobacter sp. strains GfU-1, and its close relative isolated from the same termite degraded UA only partially . Incomplete UA degradation is quite a similar phenomenon to those reported previously for isolates from the termite gut (Streptococcus sp. (Enterococcus), incomplete degradation of UA has been shown unless formate or a formicogenic substrate as a reductant is present in the medium  was acetate (73%), but formate, propionate, and butylate were also detected (10.1–4.8%). The temperature for optimum growth was 35°C and optimum pH for growth was 7.2; the specific growth rate was 0.71 μ per hour under the optimal condition. Catalase, oxidase, and urease were negative. Indol was produced. Strain NkU-1 utilized a variety of carbohydrates and produced acids from glucose, mannitol, lactose, saccharose, maltose, salicin, xylose, arabinose, gelatin, esculin, glycerol, cellobiose, mannose, melezitose, raffinose, rhamnose, and trehalose, but not from sorbitol.Clostridium saccharolyticum (98.9% identity) and Clostridium amygdalium (98.8%), although their relationships were not well resolved in the phylogenetic tree  and shows a narrower spectrum of carbohydrate utilization  is lower than that of strain NkU-1 ; therefore, strain NkU-1 likely corresponds to a novel species in the clostridia cluster XIVa, although further study, such as measuring DNA relatedness, is necessary.Based on 16S rRNA gene analysis, the closest relatives of strain NkU-1 were tic tree . C. saccflagella . C. amygReticulitermes santonensis or at a low level of UA supply (1.5 or 3.0 mM) because nitrogen of degraded UA could be first assimilated into growing cells. Taking this amount of nitrogen into consideration , entire recovery of UA nitrogen was obtained as excreted NH3 at the later stage of cell growth ; therefore, strain NkU-1 likely produced a stoichiometric amount of NH3 by UA degradation (4 mol NH3 from 1 mol UA).Addition of glucose, cellobiose, fructose, lactose, or xylose stimulated the growth of strain NkU-1 in otherwise growth-limiting medium (Y medium), but UA did not, although UA was partially degraded and NHproduced . The resproduced , and UA analyses  confirme−1) ranged from 0.90 to 0.95 and 0.11 to 0.15, respectively, after three days culture) in otherwise growth-limiting, carbon-rich but nitrogen-poor GY medium . Allantoin, allantoic acid, and ribose (ribose does not contain any nitrogen) did not increase cell growth (OD and protein (mg mL−1) reached less than 0.39 and 0.06, respectively). When ammonium chloride as a positive control was added to GY medium, OD and protein (mg mL−1) reached 0.93 and 0.12, respectively. The results suggest that strain NkU-1 utilizes various purines as a nitrogen source.In addition to UA, purine, hypoxanthine, guanine, and xanthine, and the corresponding ribonucleotides to the latter three  stimulated the cell growth of strain NkU-1 . Aerobic UA degradation generally involves uricase (urate oxidase), which needs molecular oxygen for the catalyzed reaction, and produces allantoin and allantoic acid as intermediate compounds . The anaR. flavipes and three purinolytic clostridia species; they utilize UA as a carbon and energy as well as nitrogen source and UA increases the cell yield when incorporated into growth-limiting media , necrophagy, and cannibalism likely play important roles  has been hypothesized (Blattabacterium symbiont, however, there is no candidate gene for uricolytic enzymes (Cockroaches also stored UA in their fat bodies and utilization of uric acid nitrogen by intracellular symbionts (thesized . In the  enzymes , suggest enzymes  have bee enzymes . Therefo enzymes . More exnifH, nirK, and amoA are clearly powerful tools for studies on nitrogen cycles in certain environments (e.g. Because most studies on anaerobic microbial degradation of UA were conducted more than three decades ago, modern molecular studies are anticipated. Approaches using functional key genes such as nts e.g. , 62, 63 nts e.g. , 36, 59,nts e.g. , 55 may nts e.g. , 31. The


HTLV-1/2 infection can cause severe and disabling diseases in children and adults. The aim of the study was to estimate the prevalence of HTLV-1/2 infection in pregnant women living in the metropolitan area of Rio de Janeiro.1,204 pregnant women were tested upon hospital admission for delivery in two public hospitals in the cities of Rio de Janeiro and Mesquita, between November, 2012 and April, 2013. The samples were screened by chemiluminescent microparticle immunoassay (CMIA) and reactive ones were confirmed by Western blot (WB). Epi-info software was used for building the database and performing the statistical analysis. Eight patients had confirmed HTLV-1/2 infection , equivalent to a prevalence rate of 0.66%. Two further reactive screening tests had negative Western blot results and therefore were considered negative in the statistical analysis. All HTLV-1/2-positive patients were born in Rio de Janeiro, most were non-Caucasian (87.5%), in a stable relationship (62.5%), had at least ten years of formal education (62.5%) and a monthly family income of up to US$600.00 (87.5%). There was only one case of coinfection with syphilis and none with HIV. The mean age of the infected women was 28.4 (SD = 6.3) years and of the seronegative ones was 24.8 (SD = 6.5) (p = 0.10). The median number of pregnancies were 3.0 and 1.0 (p = 0.06) and the median number of sexual partners were 3.5 and 3.0 (p = 0.33) in the seropositive and negative groups, respectively. There were no statistically significant differences between the groups.A significant prevalence of HTLV-1/2 was found in our population. The socio-epidemiological profile of carrier mothers was similar to the controls. Such findings expose the need for a public health policy of routine HTLV-1/2 screening in antenatal care, since counselling and preventive measures are the only strategies currently available to interrupt the chain of transmission and the future development of HTLV-1/2-related diseases. HTLV-1/2 are retroviruses transmitted by blood products, sexual contact and from mother to child, mainly through breastfeeding. The infection has a characteristic geographical distribution with endemic areas often neighbouring very low prevalence areas. Infection is life long and although asymptomatic in most cases, it can cause severe and disabling diseases in children and adults. There is currently no cure, vaccine or effective treatment for HTLV-1/2 infections. Our research is the first to study the prevalence of HTLV-1/2 in pregnant women living in the metropolitan area of Rio de Janeiro, the second largest in Brazil. 1,204 pregnant women were tested upon hospital admission for delivery in two public hospitals in the cities of Rio de Janeiro and Mesquita, between November, 2012 and April, 2013 and a significant prevalence of HTLV-1/2 was found (0.66%). The socio-epidemiological profile of carrier mothers was similar to the controls'. Epidemiological knowledge is fundamental for the elaboration of public health policies such as routine HTLV-1/2 screening in antenatal care, since counselling and preventive measures, mainly avoidance of breastfeeding, are the only strategies currently available to interrupt the chain of transmission and the future development of HTLV-1/2-related diseases. Human T-lymphotropic virus (HTLV)-1 and HTLV-2 were the first oncogenic human retroviruses identified in the early 1980's HTLV-1/2 infection has a worldwide distribution, with an estimate of up to 15–20 million people affected + T-cells malignancy. Type 1 virus is also associated with a wide range of inflammatory conditions, from the mild nonspecific dermatitis and uveitis to the disabling HTLV-1-associated myelopathy/Tropical Spastic Paraparesis (HAM/TSP) which affects 2–3% of the infected people Infection is life long and most of the patients remain asymptomatic, becoming viral reservoirs and keeping the chain of transmission. On the other hand, about 4% of HTLV-1 carriers will develop adult T-cell leukaemia/lymphoma (ATLL), a highly aggressive CD4There are three main routes of HTLV-1/2 transmission: sexual intercourse, infected whole blood or cell containing blood products and from mother-to-child . In endemic areas, VT remains the most important mode of transmission, since it occurs in up to 25% of the children who are breastfed by seropositive women Given its continental dimensions, Brazil is thought to be one of the countries with the highest number of HTLV-1/2 carriers in the world. Estimates range widely from 800,000 to 2.5 million people Additionally, most of the research is done on potentially biased populations such as low risk voluntary blood donors or high risk IV drug users and sex clinic attendees. Intermediate seroprevalence rates have been found in pregnant women. Although not devoid of bias, this group has been proposed to represent a more reliable portrait of the general population since their prevalence data are generally able to characterize geographic areas likely to be endemic Unfortunately there are no nationwide data on the seroprevalence of HTLV-1/2 in Brazilian pregnant women since screening is not recommended by the Ministry of Health as part of the routine antenatal care. Furthermore, there are no studies published about this prevalence on the State of Rio de Janeiro. Therefore the aim of this study was to estimate the prevalence of HTLV-1/2 infection in pregnant women admitted for delivery in the metropolitan area of Rio de Janeiro.A total of 1,204 pregnant women were recruited upon admission for delivery in two public hospitals in the metropolitan area of Rio de Janeiro, between November 2012 and April 2013. The institutions involved in the study were ‘Pedro Ernesto’ University Hospital of the Rio de Janeiro State University (Universidade do Estado do Rio de Janeiro – HUPE/UERJ), a reference for high-risk obstetric patients located in the city of Rio de Janeiro, and ‘Hospital da Mãe’, a state hospital for low and medium-complexity obstetric care, in the city of Mesquita, metropolitan area of Rio de Janeiro. Thus, both low and high-risk obstetric populations were included. Patients who did not wish to take part in the research were excluded, as well as those considered mentally unable to give consent. At the enrolment, a structured questionnaire was applied by trained interviewers to collect epidemiological, social, clinical, gynaecological and obstetric data. HTLV-1/2 positive women were counseled by a multidisciplinary team which offered information about the infection and support. Breastfeeding was contraindicated and formula milk was provided to safeguard the infants' nutrition. All the children of carrier mothers are being prospectively followed by the Paediatrics Department of HUPE.As per protocol of the Brazilian Ministry of Health all patients admitted for delivery have blood taken for HIV and Syphilis tests. At this moment, an extra sample was taken for HTLV-1/2 screening, which was performed by chemiluminescent microparticle immunoassay . Cases of reactive serology were confirmed by Western blot . All tests were carried out at the Clinical Analysis Laboratory of the HUPE/UERJ. The two patients with reactive screening tests and negative WB results were considered false positive and assigned to the negative group in the statistical analysis.This research complies with the ethical precepts established in the Declaration of Helsinki and the CONEP Resolution n° 196/96 and was approved by the Rio de Janeiro State University Research and Ethics Committee . Written informed consent was obtained from all the subjects and from the parents or guardians of the minors who agreed to take part. Anonymity and data confidentiality were guaranteed.The sample size of 1,188 patients was calculated considering an expected prevalence of 0.5% at a 95% confidence interval. Means, standard deviations, medians, interquartile ranges and percentages were used to describe the results. Medians were used to define the groups when numerical variables were analyzed as categorical ones . Missing data were excluded from the statistical analyses. Fisher Exact and Mann Whitney tests were used to assess the association between HTLV-1/2 infection and possible risk factors . Logistic regression was performed to evaluate the relation between condom use and HTLV-1/2 occurrence adjusted by age and marital status. The EPI-INFO software version 3.5.2 was used for building the database and performing the statistical analyses.Eight of the 1,204 patients had confirmed HTLV-1/2 infection (seven HTLV-1 and one HTLV-2) equivalent to a prevalence rate of 0.66%. Two further reactive screening tests had negative WB and were therefore considered false positive and excluded from the statistical analysis.In the HTLV-1/2 positive group, all the patients were born on the State of Rio de Janeiro. They were mostly non-Caucasian (7/8 - 87.5%), reported being in a stable relationship (6/8 - 62.5%), had at least ten years of formal education (6/8 - 62.5%) and 87.5% declared having a monthly family income of up to US$600.00 (approximately two minimum wages). No women reported intravenous drug use, there was only one case of co-infection with syphilis and none with HIV.vs. μ = 16.2, SD = 2.7). The median number of pregnancies were 3 and 1 (p = 0.06) and the median number of sexual partners were 3.5 and 3.0 (p = 0.33) in the seropositive and negative patients, respectively.The mean age of the infected pregnant women was 28.4 years (SD = 6.3) and of the seronegative ones was 24.8 (SD = 6.5) (p = 0.10). The mean age at first intercourse was approximately 16 years for both groups . However only a borderline statistic significance was found (p = 0.08).Antenatal groups are considered to better represent the general population than blood donors and IV drug users, being of particular interest for the chain of transmission since most of the vertical transmission can be prevented by avoiding breastfeeding.A review of the literature retrieved no published data on the prevalence of HTLV-1/2 infection in pregnant women in the State of Rio de Janeiro. The Brazilian articles concerning the infection display a significant geographic heterogeneity in prevalence, ranging from 0.07% on the State of Minas Gerais A comprehensive Brazilian study performed at the Public Blood Center Network comprising over 6 million blood donations from all the 27 state capitals revealed a striking geographic variability of the HTLV-1/2 infection The regional clustering phenomenon can be explained by the influence of the ethnical characteristics of the population in the prevalence of HTLV-1/2. The northern area of Brazil has a greater proportion of African and native-descendants than the South. There is strong evidence of endemicity of HTLV-2 among groups who originally inhabited the Amazon A strength of the present study is that all HTLV-1/2-positive subjects were born in the researched area, confirming it as a portrait of the local population. The mean age of positive pregnant women and the fact that they were mostly non-Caucasian are in accordance with the literature The similarity of the epidemiological profile in both of our groups and the lack of clear risk factors seem to point to routine HTLV-1/2 prenatal screening as the most sensible path to follow, as supported by other colleagues A limitation of our research was the possibility of selection bias. Patients were recruited at admission for delivery, which could have underestimated the overall prevalence in pregnancy since we might have lost the patients who miscarried and those who eventually had preterm labour or stillbirths.Implications for future research – our finding of such a significant prevalence shows the need for studies on a larger scale, testing all pregnant women at the beginning of antenatal care in order to avoid selection bias and increase the confidence in our results. Sampling a broader population can help to determine the cost-effectiveness of the test and justify its introduction to routine prenatal screening. Following the children of infected mothers in a cohort study could identify cases of vertical transmission and the incidence of HTLV-1/2 related diseases in our population.Universal screening for HTLV-1/2 in blood donors was implemented in Brazil in 1993 with significant impact on this mode of transmission. Additionally, horizontal transmission is thought to have been greatly reduced by successful public policies aiming at the prevention of other sexually transmitted diseases. Since 2002 HIV and syphilis prenatal screening tests are mandatory according to the Brazilian Ministry of Health in order to prevent mother-to-child transmission. HTLV screening was not included in this guideline; in fact it's not even listed as an infectious disease worthy of being targeted by government policies Since currently there is no cure, effective treatment or immunization for HTLV-1/2 infection and its complications, more accurate knowledge about its prevalence is helpful in the elaboration of public policies on educational and prophylactic measures to increase awareness and reduce the rates of viral transmission and the incidence of infection-related diseases.Supporting Information S1STROBE Checklist.(PDF)Click here for additional data file.Supporting Information S2Data file.(XLS)Click here for additional data file.Supporting Information S3Statistical analysis.(PDF)Click here for additional data file.


Adult T-cell Leukemia-Lymphoma (ATLL) is an aggressive and fatal malignancy of CD4+ T-lymphocytes associated with HTLV-1 infection, and an effective treatment is not yet available. The molecular mechanism underlying ATLL has not been fully elucidated. However, accumulation of genomic instability is believed to be a driving force for leukemogensis. How genomic instability accumulates in HTLV-1 infected cells is currently under intensive investigation. Recently, we found that the HTLV-1 viral oncoprotein, Tax, which is implicated in the chronic inflammatory response, induces DNA Double Strand Breaks (DDSB). Tax is known to activate the key T-cell inflammatory transcription factors, NF-kB, and this activation is critical for the leukemogenic process associated with HTLV-1 infection. Of note, we found that inducible nitric oxide synthase (iNOS), the enzyme that catalyzes the production of nitric oxide (NO) is highly expressed in HTLV-1 and Tax expressing cells. Interestingly, we show that the expression of iNOS is Tax-dependent and specifically requires the classical NF-kB pathway. In addition, IRF-1, the interferon regulatory factor that collaborates with NF-kB transcription factors to activate iNOS expression was also found activated by the JAK/STAT pathway. Our results show a correlation between the number of DDSB and the production of NO in tumors isolated from Tax transgenic mice. We also observed a dramatic reduction of DDSB when NO production was inhibited. Determination of the impact of NO on tumors in an ATLL mouse model will open a new area in the development of alternative strategies for the treatment/prevention of ATLL.


Ethylene gas is a major plant hormone that influences diverse processes in plant growth, development and stress responses throughout the plant life cycle. Responses to ethylene, such as fruit ripening, are significant to agriculture. The core molecular elements of the ethylene-signaling pathway have been uncovered, revealing a unique pathway that is negatively regulated. Practical applications of this knowledge can lead to substantial improvements in agriculture. In addition to being the most widely produced organic compound in the world , ethylene is a major hormone in plant biology. This volatile molecule mediates many complex aspects of plant growth, development and survival throughout the plant life cycle, including seed germination, root development, shoot and root growth, formation of adventitious roots, abscission of leaves and fruits, flowering, sex determination, and senescence of flowers and leaves [The simple hydrocarbon ethylene  . EthylenPlants synthesize ethylene using a two-step biochemical pathway starting from S-adenosyl-L-methionine (SAM) , 6 Fig.. SAM is Ethylene is biologically active at very low concentrations of around 0.01 to 1.0 part per million (ppm). Lower or higher sensitivities have been observed depending on the species and the response. Some climacteric fruits, such as tomatoes and apples, can generate tens of ppm of ethylene. It is worth noting here that ethylene is a byproduct of partial combustion of organic fuels and is present, therefore, in the atmosphere due to such things as forest fires, volcanic eruptions and car exhaust .Metazoans lack ethylene receptor homologs and do not perceive and respond to ethylene as plants do. High concentrations of ethylene (>1000 ppm) can, however, cause dizziness or light-headedness. For several decades in the 1900s, ethylene was used as a general anesthetic . In anciArabidopsis thaliana, initiated by the isolation of Arabidopsis ethylene response mutants. Mutants were isolated in the late 1980s concomitant with the development of Arabidopsis as a genetic plant model [Arabidopsis seedlings exhibit a short and thick hypocotyl, an exaggerated apical hook and a short root  andy tomato .Fig. 2ThEthylene signaling involves a unique pathway, shown in Fig. Arabidopsis has five ethylene receptors and tomato has six) that have overlapping and distinct functions [The ethylene receptor is unexpectedly related to the histidine protein kinase receptors of the two-component signaling system, which is prevalent in prokaryotes but very rare in eukaryotes. It is believed that plants most likely acquired the ethylene receptor gene from an ancient endosymbiotic cyanobacterium that became the chloroplast . Plants unctions , 17. As The ethylene receptors form disulfide-linked dimers, and each dimer is capable of binding a single ethylene molecule  with thectr1 loss-of-function mutants, whereas dominant, gain-of-function receptor mutations confer ethylene insensitivity [This is a good question. Currently this is best understood at the genetic level. From genetic analyses, we know that the receptors are negative regulators of ethylene responses. In other words, ethylene responses are repressed by ethylene receptor signaling . This resitivity .This is still unresolved. In the canonical two-component system, binding of the ligand either stimulates or inhibits autophosphorylation of a conserved histidine residue followed by transfer of the phosphate to a conserved aspartate in the receiver domain. Curiously, histidine kinase activity does not appear to play a major role in ethylene receptor signaling , 17. AltAlthough genetic evidence indicates that ethylene binding inhibits receptor signaling, there is no clear answer to the basic question: Does the binding of ethylene stimulate or inhibit biochemical activity in the receptor? There are data to support each possibility. Although it might be counter-intuitive, a formal possibility is that CTR1 activation occurs by a passive  signaling mechanism that is alleviated when ethylene receptor activity is triggered by the binding of ethylene. Complete structural data for the receptors, which is not yet available, will hopefully shed light on this question. It is also unclear why plants have multiple ethylene receptor isoforms. Although there is evidence that individual receptors have distinct roles in controlling specific responses, the underlying mechanisms for sub-functionalization are unknown .EBF1/2 mRNA [EBF1/EBF2 ubiquitin-ligating complex [Ethylene signaling downstream of CTR1 hinges on the phosphorylation status of EIN2, an enigmatic central regulator of the ethylene-signaling pathway . EIN2 is1/2 mRNA , 27. Thi1/2 mRNA . In the  complex ; this isAPETELA2 (AP2)/ETHYLENE RESPONSE FACTOR (ERF) family, such as ERF1, which regulates further expression in a transcriptional cascade of ethylene signaling [EIN3 was shown to initiate a transcriptional cascade that triggers several dynamic waves of gene expression that include feedback loops and the activation of genes known in numerous other hormone signaling pathways . The priignaling , 30. Theignaling .ein2 mutations have been obtained in genetic screens for components of other signaling pathways [Given that plant growth, development and stress responses require the integration of diverse environmental and endogenous signals, there is a growing focus on ethylene cross-talk with other signals , 32. Mulpathways , EIN2’s etr1-1 gene of Arabidopsis [ERF transcription factor gene (SUB1A) [Continued advances in understanding cross-talk and transcriptional networks will lead to a deeper understanding of ethylene-signaling networks that will someday allow for the modeling of specific plant responses. Applications of such knowledge have tremendous potential for agricultural improvements. Already, our understanding of ethylene biology can provide new strategies for manipulating ethylene responses, particularly by genetic means. Examples include the delay of flower senescence by expressing a dominant mutant bidopsis  and the  SUB1A) . The dayA . The d


Human T-cell Lymphotropic Virus type 1 (HTLV-1) is a human retrovirus that infects at least 5–10 million people worldwide, and is the etiological agent of a lymphoproliferative malignancy; Adult T-cell Leukemia/Lymphoma (ATLL); and a chronic neuromyelopathy, HTLV-1 Associated Myelopathy/Tropical Spastic Paraparesis (HAM/TSP), as well as other inflammatory diseases such as infective dermatitis and uveitis. Besides sexual intercourse and intravenous transmission, HTLV-1 can also be transmitted from infected mother to child during prolonged breastfeeding. Some characteristics that are linked to mother-to-child transmission (MTCT) of HTLV-1, such as the role of proviral load, antibody titer of the infected mother, and duration of breastfeeding, have been elucidated; however, most of the mechanisms underlying HTLV-1 transmission during breast feeding remain largely unknown, such as the sites of infection and cellular targets as well as the role of milk factors. The present review focuses on the latest findings and current opinions and perspectives on MTCT of HTLV-1. Diffethe host . In eachthe host . It is net al. [et al. [First evidence of HTLV-1 transmission from infected mother to children during lactation has been brought by epidemiological studies. HTLV-1 infection was more prevalent among breastfed children than bottle-fed children in Japan ,9, with et al.  reported [et al.  showed t [et al. ). HTLV-1 [et al. . The maj [et al. . Of note [et al. ,17,18.From a virological point of view, it is known that many retroviruses may be transmitted via breast milk, such as Moloney murine leukemia virus ,20, MousCallithrix jacus) was able to induce seroconversion within 2.5 months, and the virus was detected in the animal peripheral blood lymphocytes [7 cells from ATLL patients were sufficient to infect these animals. Similarly, oral inoculation of concentrated fresh milk from HTLV-1 seropositive mothers in the same animal model could transmit the infection [i.e., MT-2) to rats induced a persistent HTLV-1 infection in the absence of both humoral and cellular immune responses [From an experimental point of view, oral inoculation of peripheral blood lymphocytes isolated from ATLL patients to adult common marmosets  on oral inoculation of HTLV-1 were mainly focused on seroconversion, progression towards an ATLL-like disease, and immune status of the host. Among the different possible sites of entry, the palatine tonsils and gut seem to be of particular importance due to their enrichment in possible targets (lymphoid cells and M cells), their function in antigen sampling  and theiin vitro studies have been published concerning the mechanisms of passage of HTLV-1 across the intestinal barrier. In 1992, Zacharopoulos et al. [i.e., I407) was susceptible to HTLV-1 infection in vitro, as shown by electron microscopy, in situ hybridization, and PCR amplification. However, it was unclear if the cells were fully differentiated as no data on the epithelium integrity and tight junctions were shown. In contrast, our laboratory performed studies on the susceptibility of three different human enterocytic cell lines on compartmentalized culture devices, with assessment of the enterocyte differentiation: existence of a tight epithelial barrier was checked by electron and confocal microscopy, and the trans-epithelial resistance was measured [Very few s et al.  indicatemeasured . In thismeasured ,50. Howemeasured .in vitro studies [HTLV-1-infected lymphocytes, once they have crossed the mucus layer, could either pass through the epithelium via M cells, or in-between enterocytes (paracellular passage), to reach the lamina propria where potential targets of HTLV-1 infection, such as T lymphocytes (and/or dendritic cells and B lymphocytes) are located. From  studies , the infin vitro results highlight one of the potential mechanisms proposed for HTLV-1 passage, which are summarized in Interestingly, viral transcytosis through enterocytes was observed only in the presence of HTLV-1 infected lymphocytes, and not in the case of purified virions. Such a mechanism of viral transcytosis is reminiscent of previous work showing transcytosis of HIV across an epithelial barrier, without infection of enterocytes, and subsequent infection of macrophages or CD4 lymphocytes located to the basal side of the epithelium . These iStudies on HTLV-1 MTCT determinants have focused mainly on genetic host factors, immunological host factors, lactation duration and milk components.et al. [HLA (Human Leukocyte Antigen) genes distribution was different for ATLL or HAM/TSP patients compared to asymptomatic carriers , but th [et al.  indicateome 6q27 .in vitro that the addition of HTLV-1 serum cord blood plasma is able to prevent the infection of human neonatal lymphocytes when co-cultured with breast-milk cells of HTLV-1 carrier mothers [Concerning the immunological factors involved in HTLV-1 MTCT, the role of maternal anti-HTLV-1 antibodies may appear controversial. Such studies have to take into account the duration of breastfeeding, since the protective role of anti-HTLV-1 antibodies has been demonstrated in a rabbit model of infection, where passive immunization was shown to prevent milk-borne transmission of HTLV-1 to offspring . Moreove mothers . However mothers ,57,58,59 mothers .Another point to take into account concerns the other milk components that may influence HTLV-1 MTCT transmission. As an example, it has been shown that lactoferrin, an iron-binding milk glycoprotein, was able to enhance HTLV-1 replication, by transcriptional activation of HTLV-1 Long Terminal Repeat (LTR), the viral promoter . This efin vivo using a humanized mouse as a model of HTLV-1 infection [in vitro approach, such as differentiation of M cells from enterocytic cell lines, on compartmentalized culture devices, as already done to show the role of these cells in virus transport across the epithelium [in vivo/in vitro studies will also allow delineating the role of factors such as milk components , the proviral load, and the antibody titer in HTLV-1 transport through the intestinal epithelium.One of the major remaining questions on MTCT concerns the sites of primary passage/infection of HTLV-1 in the digestive tract. The mechanisms of HTLV-1 infection after oral inoculation should be addressed nfection ,64,65, iithelium . CombinaAnother perspective concerns the comprehension of the role of breastfeeding duration on HTLV-1 MTCT. It seems rather clear that such a role corresponds to a combination of the cumulative viral input, the changes over time in milk composition in infected cell types, maternal antibodies, and the immune status and maturation of the neonate’s gut. It is known that neonatal life  organizes and controls mucosal homeostasis through endogenous and exogenous factors that drive the development and maturation of the intestinal immune system . RecentAltogether, these further studies could delineate new preventive strategies to counteract HTLV-1 MTCT, as well as provide new data on the general mechanisms of pathogenic agents in MTCT.


Uric acid is the end product of purine metabolism in humans, and increased serum uric acid concentrations lead to gout. The objective of the current study was to identify factors that are independently associated with serum uric acid concentrations in a cohort of Czech control individuals.MTHFR , SLC2A9  and ABCG2 genes (c.421C>A). A backward model selection procedure was used to build two multiple linear regression models; in the second model, the number of metabolic syndrome criteria that were met replaced the metabolic syndrome-related variables.The cohort consisted of 589 healthy subjects aged 18–65 years. We studied the associations between the serum uric acid concentration and the following: (i) demographic, anthropometric and other variables previously reported to be associated with serum uric acid concentrations; (ii) the presence of metabolic syndrome and the levels of metabolic syndrome components; and (iii) selected genetic variants of the ABCG2, unlike MTHFR, was a genetic determinant of serum uric acid concentrations.The models had coefficients of determination of 0.59 and 0.53. The serum uric acid concentration strongly correlated with conventional determinants including male sex, and with metabolic syndrome-related variables. In the simplified second model, the serum uric acid concentration positively correlated with the number of metabolic syndrome criteria that were met, and this model retained the explanatory power of the first model. Moderate wine drinking did not increase serum uric acid concentrations, and the urate transporter ABCG gene are independently associated with the serum uric acid concentration. Our model indicates that uric acid should be clinically monitored in persons with metabolic syndrome.Metabolic syndrome, moderate wine drinking and the c.421C>A variant in the  An increased serum uric acid (UA) concentration (>416 µmol/L in men and >340 µmol/L in children and women) is an important risk factor for gout and has other significant associations with human disorders, such as cardiovascular and renal diseases UA is the end product of purine metabolism in humans and higher primates; in contrast, other mammals further metabolize urate to allantoin. The absence of the hepatic enzyme uricase (urate oxidase) and efficient renal UA reabsorption contributes to the ten-fold higher UA blood levels in humans compared to those of other mammals. UA is a powerful scavenger of peroxyl radicals, hydroxyl radicals and singlet oxygen in human biological fluids. The balance between UA production and excretion determines serum UA concentrations; the daily production of urate is approximately 1000 mg (6 mmol) in adults. Approximately 75% of UA excretion occurs via the kidney, and the gastrointestinal tract eliminates 25%. The renal excretion of UA depends on the balance between filtration, secretion and reabsorption. Transporters for urate localize in the proximal tubule, where specific transporter proteins in the apical and basolateral membranes secrete and extensively reabsorb urate. As a result, approximately 10% of the UA filtered through the glomerular membranes is excreted.Many studies have shown an association between alcohol consumption and hyperuricemia, which is a common precipitating cause of acute gouty arthritis SLC2A9, SLC22A12 and ABCG2 genes. The URAT1 transporter, the product of SLC22A12, was identified in 2002 as the crucial transporter involved in UA reabsorption. The URAT1 transporter regulates blood UA levels and plays a central role in the reabsorption of UA from the glomerular filtrate SLC22A12 gene cause renal hypouricemia type 1, OMIM #220150 SLC2A9 in UA reabsorption was identified SLC2A9 and serum UA levels, the excreted fraction of UA, blood pressure, body mass index and gout SLC2A9 genetic variants are responsible for a portion of the variance in serum UA concentrations: 5–6% in females and 1–2% in males SLC2A9 gene cause renal hypouricemia type 2, OMIM #612076 ABCG2 and causes approximately 10% of the hyperuricemia and gout cases in Caucasians Over the past decade, genome-wide association studies and meta-analyses have led to a massive increase in our knowledge of the common genetic variants that influence serum UA concentrations. However, detailed knowledge of the degree to which genetic variants predict serum UA concentrations and of the functional pathway through which these variants may alter serum UA remains limited. The genes that influence the level of serum UA via renal UA excretion are those for the urate transporters, primarily those encoded by the MTHFR, 1p36.22, EC 1.5.1.20) and the serum UA concentration was also reported MTHFR allelic variants may be confounding genetic factors in the relationship between serum UA and hypertension Although most genes associated with UA are involved in the renal urate transport system, an association between homozygosity for the common genetic variant c.665C>T  of methylenetetrahydrofolate reductase  What is the relationship between serum UA and demographic, anthropometric and other variables? (b) What is the relationship between serum UA and metabolic syndrome? (c) What is the influence of genetic determinants on serum UA? In our study, we examined the associations between serum UA concentrations and a wide range of variables in a cohort from central Europe. We modeled the associations between UA and (i) demographic, anthropometric and other variables previously reported to be associated with serum UA ; (ii) the presence of metabolic syndrome (MS) and the levels of MS components and (iii) selected genetic variants in the SLC2A9, ABCG2 and MTHFR genes. We have adopted a complex approach, combining all these variables into one regression model.Even with extensive research We have used the data and genomic DNA samples from subjects of a previously described study 2 and/or a WHR ≥0.90 for men and ≥0.85 for women UA in the serum was measured using a uricase enzymatic method by an auto-analyzer. We studied the associations between the serum UA concentration and the following demographic characteristics, metabolite levels, conventional risk factors and genetic risk factors: (i) conventional variables—sex, age, alcohol consumption (beer and wine), smoking status and number of packyears, allopurinol use, diuretic use, acetylsalicylic acid consumption, hormonal treatment, hormonal contraception and serum creatinine levels, (ii) metabolic syndrome-related variables—BMI, WHR (waist-to-hip ratio), systolic and diastolic pressure, hypertension, treatment of hypertension, hyperglycemia, diabetes mellitus and its treatment, total cholesterol, HDL and LDL cholesterol, triacylglycerol levels, hyperlipidemia and the gamma-glutamyltransferase (GGT) concentration and (iii) selected genetic variants (see above). For the second analysis, we constructed a new variable called metabolic syndrome (the number of metabolic criteria met) and used it instead of the MS-related variables. We used the following common criteria for MS reported by several organizations, including the WHO, the IDF and the AHA/NHLBI SLC2A9  and c.881G>A ), the urate transporter ABCG2 ) and the MTHFR enzyme  and c.1286A>C ) were selected for genotype analysis. The selected genetic variants of the urate transporters were analyzed by PCR with allele-specific primers (ARMS-PCR); the primers and conditions are given in MTHFR variants were determined in a previous study The frequent genetic variants of genes encoding the urate transporter In total, 30 potential predictors were included in the analysis. For statistical purposes, the UA concentration and the glucose, GGT and triglyceride levels were logarithmically transformed. The association between the variables and the serum UA concentration was evaluated in two steps. First, single-predictor linear regression models were utilized with the UA concentration as the response variable and one predictor variable adjusted for sex (including possible interactions with sex). Then, the backward model selection procedure was used to build a multiple linear regression model. This model was based on the set of variables that showed association  in the single-predictor models, together with all interaction terms with sex. In the second multiple linear regression analysis, we started with the same large set of variables in interaction with sex, but we replaced the predictor variables related to metabolic syndrome with the number of MS criteria met and repeated the backward selection procedure.2 value. The logarithmic transformation of the UA concentration made it difficult to interpret the values of the coefficients in the multiple regression models. We therefore calculated the model-predicted mean change in UA concentration when the studied variable increased by one unit while the values of the other variables remained the same for an individual whose original UA concentration was 326 µmol/L .We assessed the relative effect of the genetic component in the final regression models by analyzing the proportion of the explained variance relative to the adjusted Rhttp://www.R-project.org).The level of statistical significance was set to 0.05 for the coefficients in the multiple regression analysis models. All analyses were performed in the statistical language and environment R, version 2.14.0 ; since the genotyping errors are unlikely this observation may be due to the population stratification or by random occurrence. There was a significant additive effect of ABCG2 c.421C>A  on the increase in the UA concentration. The single-predictor analysis of the effect of the SLC2A9 c.881G>A variant showed a possible additive effect on the decrease in the serum UA concentration , whereas the SLC2A9 c.844G>A variant did not show any effect on the UA concentration. No significant relationship was found between the MTHFR variant c.665C>T and the UA concentration. Homozygous individuals with the MTHFR c.1286A>C variant had significantly lower UA concentrations  than those of the remainder of the study population. However, when MTHFR c.1286A>C was considered as a three-level factor , the model p-value was greater than the significance level .Genetic factors were significantly associated with the serum UA concentrations. The allelic and genotype frequencies and geometric means of serum UA concentrations in wild-type homozygotes, heterozygotes and variant homozygotes for different loci are shown in In the study population, 30% of the subjects had MS (106 men and 70 women met three or more MS criteria), and 70% of the subjects did not have MS (178 men and 233 women). The box and whisker plot in MTHFR c.1286A>C, ABCG2 c.421C>A, SLC2A9 c.844G>A and SLC2A9 c.881G>A. The adjusted coefficient of determination (R2) was 0.59. The genetic variables together explained approximately 4.5% of the total variability.To assess the independent contributions of the variables to the UA concentration, a multiple linear regression model was built . The SLC2A9 c.881G>A and c.844G>A variants showed negative effects on the serum UA concentrations. In both models, the serum UA concentration was lower for individuals with the MTHFR c.1286CC genotype.In both models, the serum UA concentration was higher for the individuals with the C allele of the a) the serum UA concentration was positively correlated with the number of MS criteria that were met, and moreover, this simplification had no substantial influence on the explanatory power of the model; (b) moderate wine drinking likely does not increase serum UA concentrations; and (c) the urate transporter ABCG2, unlike MTHFR, is one of the genetic determinants of serum UA concentrations. The evidence for an influence of two SLC2A9 gene variants is inconclusive.In the present study, we demonstrated the following new findings: . Several studies explored the links between increased serum UA concentration and various components of MS vice versa and for screening for hyperuricemia-related problems in patients with MS in clinical practice.We showed that the serum UA concentration correlates positively (p<0.001) with the number of MS criteria  and was independent of all other covariates. Our results confirm that this alteration in the ABCG2 gene is a common cause of hyperuricemia due to decreased renal urate excretion. This finding is consistent with previously reported studies in which the ABCG2 c.421C>A variant SLC2A9 c.844G>A and c.881G>A variants were associated with a significant decrease in serum UA concentrations. This result is surprising, as all other studies reported increased serum UA levels or mixed results in individuals carrying c.844G>A and c.881G>A SLC2A9 was associated with elevated serum uric acid concentration  in the Framingham and Rotterdam cohorts SLC2A9 gene was significantly associated with elevated serum uric acid concentrations and gout in the Han Chinese, Solomon Island and Japanese cohorts SLC2A9 variants with serum UA concentrations in the Czech cohort.Our study showed a strong association between the allelic variant c.421C>A in the Using a multiple linear regression model, we confirmed the previously reported sex-specific differences and the strong associations between serum UA concentration and age, menopause, BMI, hypertension, alcohol consumption, serum triacylglycerol levels, creatinine levels and GGT levels Our study has several strengths. First, we explored the association between serum UA concentrations and several important demographic, anamnestic, biochemical and genetic factors in a Central European (Czech) population with detailed characterization that allowed us to answer our research question and that could be generalized to the larger population. Second, we controlled for several potential confounders, such as the types of medications, that may influence serum uric acid concentrations. Third, we used a complex approach, a multiple linear regression model that provides a comprehensive estimate of the association. Different limitations of this study should also be acknowledged. First, the size of the cohort may not be sufficiently large for such detailed analysis. Second, the number of the frequent genetic variants of genes encoding the urate transporters was limited.ABCG2 is one of the genetic determinants of serum UA concentrations. These data may have implications for the clinical prediction of UA concentrations and may lead to personalized medical diagnoses and treatments for common diseases. An increased serum UA concentration is significantly associated with MS. In clinical practice, hyperuricemia is an indication for investigating MS criteria, and the presence of MS is an indication for investigating the serum UA concentration.In conclusion, we showed that the serum UA concentration positively correlated with the number of MS criteria that were met. We found that moderate wine drinking may not increase serum UA concentrations. We confirmed that the urate transporter Table S1Genotype analysis - arrangement and primers. The selected genotypes were determined using multiplex ARMS-PCR. The amplifications were performed in the total volume 10 µl using Combi PPP master mix  supplied with allele specific primer pairs as it is shown in this table. The cycling conditions were  in DNA Engine Dyad PTC-220 . PCR products were analysed on 3% agarose gels.(DOC)Click here for additional data file.Table S2The final multiple linear regression model (Model 1) and the alternative model, with MS predictors replaced by graded MS (Model 2). * log-transformed for the model, coefficient and standard error relevant for the transformed variable. ** values for man/woman because of an interaction with gender present in the model.(DOC)Click here for additional data file.


Zea mays), barley (Hordeum vulgare), and soybean (Glycine max) yields. Fertilizer contributed the most to the barley yield and FYM contributed the most to the soybean yield among the three crops. The baseline fertility of the subsurface soil was very low for maize and barley prior to fertilization. In contrast, the baseline fertility in this soil approximated half-saturated fertility for the soybean crop. The long-term soil fertility was increased by adding FYM, but the effect of FYM addition was reduced by the addition of fertilizer. Our results provide evidence that long-term soil fertility under continuous farming was maintained, or increased, by the application of natural nutrients compared with the application of synthetic fertilizer.To effectively manage soil fertility, knowledge is needed of how a crop uses nutrients from fertilizer applied to the soil. Soil quality is a combination of biological, chemical and physical properties and is hard to assess directly because of collective and multiple functional effects. In this paper, we focus on the application of these concepts to agriculture. We define the baseline fertility of soil as the level of fertility that a crop can acquire for growth from the soil. With this strict definition, we propose a new crop yield-fertility model that enables quantification of the process of improving baseline fertility and the effects of treatments solely from the time series of crop yields. The model was modified from Michaelis-Menten kinetics and measured the additional effects of the treatments given the baseline fertility. Using more than 30 years of experimental data, we used the Bayesian framework to estimate the improvements in baseline fertility and the effects of fertilizer and farmyard manure (FYM) on maize ( Agricultural crop production is directly related to food supply, so agricultural soil productivity must be maintained. Balanced fertilization provides all the essential nutrients for crops to remain healthy and grow productively The relationship between crop yield and soil fertility under different fertilization regimes has been studied for decades. Aref and Wander Many researchers have suggested that soil fertility under continuous farming is maintained, or increased, by the application of farmyard manure (FYM) compared with the application of synthetic fertilizer Previous models have predicted the crop-yield response to fertilizer application Zea mays), barley (Hordeum vulgare), and soybean (Glycine max) yield, we estimated in the Bayesian framework the improvements in baseline fertility and the effects of fertilizer and farmyard manure (FYM). We compared the efficiency of separate applications of fertilizer or FYM to the three crops  using estimated model parameters. The temporal variations in the baseline fertility of each crop were estimated and compared for six treatments and two soil types.Soil quality is a complex combination of biological, chemical, and physical properties. In this paper, we focus on the application of these concepts to agriculture and define the baseline fertility as the level of soil fertility that crops utilize for their growth. Therefore, our soil quality is an interaction of the above properties and the crop activities. With this strict definition of baseline fertility, we propose a new crop yield-fertility model that enables quantification of the process of improving baseline fertility and the effects of treatments solely from the time series of crop yields. The model was modified from Michaelis-Menten kinetics and measured the additional effects of the treatments given the baseline fertility. Using more than 30 years of experimental data of maize . The field site was located in the Kantō Plain, where the soil is covered with pyroclastic material from volcanoes that surrounded the western Kantō region 126,000 years ago. The soil parent material is Tachikawa loam. The surface soil is a black-colored fertile andosol containing a high percentage of humus and the subsurface soil (25 cm below the surface) is red-colored barren clay. Andosols are soils found in volcanic areas formed in volcanic tephra. Andosols have a different composition from chernozems and are not commonly found outside the Pacific “ring of fire”. At the beginning of the experiment, the percent nitrogen was 2.23 g/kg in the surface soil and 0.97 g/kg in the subsurface soil. A 2-year crop rotation of maize-barley-soybean-barley was maintained throughout the experimental period and the crop yields were measured. The maize crop was sown at the beginning of July and harvested at the end of September every second year from 1980 onwards. The barley crop was sown in the first half of November and harvested either at the end of May or at the beginning of June of the next year each year from 1980 onwards. The soybean crop was sown at the end of June and harvested in the second half of October every second year from 1981 onwards. The seeds of the three crops were ridge sown using a seeding machine. For maize and soybean, one seed was sown in each hole, and the widths between the ridges and stocks were 71 cm and 23 cm, and 71 cm and 17.5 cm, respectively. For barley, row seeding was applied using a width of 17.8 cm between the ridges.2 per plot, 8×8 m) and the subsurface soil  and were replicated four times. The percent nitrogen, phosphorus, and potassium in the fertilizer treatments were N:P:K = 12:7.9:13.3 for maize and barley and 3:4.4:8.3 for soybean. Fertilizer was added to the soil at a rate of 1 t ha−1 for all crops. The FYM comprised wheat straw and cow dung. The percent nitrogen in the FYM ranged from 0.2–0.4. FYM was added to the plots at two levels, i.e., 20 and 60 t ha−1. The FYM treatments were added to the soil twice a year from 1980 to 1990 before the crops were sown and once a year (before the barley was sown) from 1991 onwards. After the barley harvest in 2008, fertilizer and FYM applications were discontinued for all treatments. Phosphate fertilizer was added simultaneously with the NPK fertilizer and FYM at a rate of 2 t ha−1 after the soybean harvest from 1980 until 2001. All aboveground components were removed from the field during harvesting. The dead foliage, below-ground components, and remnant stem sections  remained in the field. The harvested material from the six treatments for both the surface and subsurface soils were weighed after drying at 80°C. The dry weight (g m-2) of the aboveground maize components, the barley spike, and the soybean seed were measured.Six treatment plots containing the NPK fertilizer applications and two levels of farmyard manure (FYM) combined with compost were established in both of the fields with the surface soil . Although the crops were planted in the same fields with the same soil, they obtained and used nutrients from the soil in a manner unique to each crop.The crop yields of the surface and subsurface soils without the addition of fertilizer or farmyard manure (FYM) reflect the yield differences that occurred purely as a result of the soil. The crop yields observed for the subsurface soil were 39±25%, 49±20%, and 19±31% (±SD) lower than the yields observed for the surface soil for maize , barley P>0.2) between the surface and subsurface soils when fertilizer or FYM or a combination of fertilizer and FYM were applied (P<0.05) or FYM alone  were added, but did not differ significantly when only fertilizer was added  (The maize and soybean yields did not differ significantly ( = 0.52) . The difP<0.05) for both soil types over the 30-year period. The barley yield increased significantly in the subsurface soil (P<0.05) but not in the surface soil (P = 0.15). The soils were fertilized by crop residues ) that remained in and on the soil after harvesting. Organic matter accumulated in the soil over time, providing a long-term, slow-release source of nitrogen, phosphorus, sulfur, and other important nutrients for crop growth, which resulted in increased soil productivity.The maize and soybean yields increased significantly (P<0.001). Maize and soybean yields  did not differ significantly between the two soil types. The interaction effects of fertilizer and FYM were significant for maize  and barley , but not for soybean . The soybean yield did not differ significantly among the treatments (P>0.1 for both fertilizer and FYM).We used a nested analysis of variance to compare the yields of the three crops in the surface and subsurface soils in response to the different treatments. The surface soil produced significantly greater barley crop yields compared with the subsurface soil  to the total fertility (i at time t (BF) for the subsurface soil without the addition of fertilizer or FYM in the first year was normalized by setting this value to 1. V is the maximum yield in response to the maximum fertilizer input; K is the fertility level before fertilization that is required to produce half of the maximum yield, V. A large K indicates that more nutrients need to be added to the soil for the crop to grow, so K can infer the sterility of the soil prior to fertilization. i at time t and is assumed to vary gradually over time; F is the level of fertilizer application (0 and 1), M is the level of FYM application , and a and b represent the contributions of F and M relative to the baseline fertility, respectively. Hereafter, we refer to V, K, a, and b as the maximum yield, half-saturated fertility, fertilizer contribution, and FYM, respectively. A Bayesian framework was adopted for parameter estimation assuming a gradual change in the baseline fertility.We did not apply the model to describe the kinetics. Rather, we used a form of the model to interpret the effects of baseline fertility, fertilizer, and FYM on the crop yield and the extent of saturation of additional inputs. The model related crop yields  were increased by continuous cropping over the experimental period. The difference in the baseline fertility between the treatments was 0.89 in 1980 and 3.24 in 2008 for maize; for barley, it was 1.20 in 1980 and 2.54 in 2010, and for soybean, it was 0.96 in 1980 and 3.19 in 2009.P<0.001) and barley , but not for soybean . The baseline fertility of soybean differed significantly for fertilizer  but not for FYM .We used a 2×2 factorial analysis of variance to compare the baseline fertility of the three crops in response to the different treatments. The interaction effects of fertilizer and FYM were significant for maize  compared with barley and soybean. Fertilizer contributed the maximum amount to the barley yield, and the FYM contributed the maximum amount to the soybean yield.The total fertility levels among the treatments for both surface and subsurface soils during the 30-year period ranged from 0.54 to 7.22 for maize , 0.41 to−2 for maize, 107.70 to 764.56 g m−2 for barley, and 117.02 to 269.52 g m−2 for soybean. Because of fluctuations in crop yield that could not be described by the model, the predicted yields covered 49, 57, and 42% of the observed yield ranges for maize (35 and 2023 g m−2), barley (14 and 1168 g m−2), and soybean (16 and 381 g m−2), respectively.Using the total fertility range, we estimated the predicted range in crop yield using the crop yield-fertility model for each crop. The predicted crop yields ranged from 236.24 to 1217.51 g mIn the sterile subsurface soil in the initial year, the application of fertilizer resulted in 153, 189, and 62% increases  in crop yield for maize, barley, and soybean, respectively. In the fertile surface soil in the initial year, the application of fertilizer resulted in 275, 196, and 35% increases  in crop yield for maize, barley, and soybean, respectively. After 20 years of farming, the application of fertilizer resulted in 89, 219, and 11% increases  in the subsurface soil, and 62, 113, and 7% increases  in the surface soil for maize, barley, and soybean, respectively, because of the increased baseline fertility. The greatest yield increases were observed in the initial year in the fertile surface soil with the application of both fertilizer and FYM (compared with the treatment without fertilizer or FYM), and then the yield increment decreased each year . In the P<0.001) for maize, 0.801 (P<0.001) for barley, and 0.622 (P<0.001) for soybean.The estimated fertility levels were used to predict the crop yield using the yield-fertility curves for each crop , and S4Ae, 0.801 , P<0.001nd 0.622 , P<0.001Yield fluctuations were observed among the treatments but these fluctuations were not reflected by the predicted crop yields. For maize, the peak yields occurred in 1990 and 2004, and a reduced yield occurred in 1996. For barley, the peak yield for the fertilizer-treated plots occurred in 1993, but the peak yield occurred 1 year later in the plots without fertilizer treatment. The FYM treatments were discontinued in 1991 for the summer harvested crops (maize and soybean). Clear reductions in maize yield in 1991 and soybean yield in 1992 are shown in Crop yield can be affected by climatic factors The baseline fertility of the subsurface soil was very low for maize and barley. This suggests that the fertility in the subsurface soil prior to the application of fertilizer or FYM was not sufficient for maize or barley.The increasing baseline fertility trend was also observed in soil without additional fertilizer or FYM. The increased fertility may have resulted from the addition of crop residues to the soil and nitrogen fixation by the soybean crop. These fertility increases may indicate soil maturation  in the experimental fields.The baseline fertility of maize indicated that when FYM was applied alone, soil fertility increased more than when both FYM and fertilizer were applied, for both soil types . The appOur result indicates that the amount of fertilizer applied in the experiment reached the fertility level at which the maize yield was half of the maximum yield, but the amount of FYM required to reach this value was much lower. Barley obtained the highest fertility from the soil and fertilization despite the fact that maize produced the greatest crop yield and had the largest half-saturated fertility level. By using nutrients from the soil and fertilizer, soybean was the crop that most often approached the maximum yield.Our findings indicate that the sterile subsurface soil needed more time to accumulate nutrients than the surface soil. The percentage increments in crop yield were larger in the subsurface soil than in the surface soil, which indicates that fertilizer application effectively improved the crop yield in the barren field. The fertilizer-induced increment in maize and barley crop yield increased after 2001. This is because the phosphate fertilizer application was discontinued in 2001 and the baseline soil fertility subsequently declined. The soybean crop did not show a similar reduction in yield after 2001.Long-term experiments examining fertilizer treatments and crop yield have been widely conducted to assess the effects of applying fertilizer By allowing the gradual temporal variation in baseline fertility, it was possible to estimate the process of soil fertilization and the short-term effects of different fertilizer treatments. Our results support the proposal that naturally derived nutrients should be used to maintain soil fertility and synthetic fertilizer should be used to maintain productivity. Our results provide a clear description of the relationship between crop yield and soil, which can be easily understood.Figure S1−2) for six treatments in a field with a fertile surface soil and a field with a barren subsurface soil from 1980 to 2010.Temporal variations in observed crop yield (g m (A) Maize; (B) Barley; (C) Soybean; SS, subsurface soil; S, surface soil; F, level of fertilizer; M, level of farmyard manure.(TIF)Click here for additional data file.Figure S2−2) of maize for six treatments in a field with a fertile surface soil and a field with a barren subsurface soil presented every second year from 1980 to 2008.Temporal variations in the observed and predicted yields (g m (A) A crop yield-fertility map. The crop yield-fertility model translates total fertility into predicted yield. The gray band represents the band that corresponds to the standard deviation (±SD) of the crop yield-fertility curve. (B) The relationship between the observed and predicted yields. SS, subsurface soil; S, surface soil; F, level of fertilizer; M, level of farmyard manure; BF, baseline fertility.(TIF)Click here for additional data file.Figure S3As for (TIF)Click here for additional data file.Figure S4As for (TIF)Click here for additional data file.Figure S5Temporal variations in the baseline fertility estimated using the crop yield-fertility model for six treatments in a field with a fertile surface soil and a field with a barren subsurface soil from 1980 to 2010. (A) Maize; (B) Barley; (C) Soybean; SS, subsurface soil; S, surface soil; F, level of fertilizer; M, level of farmyard manure.(TIF)Click here for additional data file.Figure S6Temporal variations in the posterior means and the bands that correspond to the standard deviation (±SD) of the Bayesian estimates of baseline fertility for six treatments in a field with a fertile surface soil and a field with a barren subsurface soil. (A) Maize; (B) Barley; (C) Soybean; SS, subsurface soil; S, surface soil; F, level of fertilizer; M, level of farmyard manure.(TIF)Click here for additional data file.Figure S7Traces of the MCMC samples of V−2),   Maize; (B) Barley; (C) Soybean. The chain length was set to 1,000,000 steps logging every 100th step.(TIF)Click here for additional data file.Figure S8Temporal variation of the climatic variables recorded at the experimental site from January 1980 to December 2011. (A) Monthly precipitation. (B) Average monthly temperature.(TIF)Click here for additional data file.File S1Contribution of the climatic factor. The analyses by including a temperature effect in the crop yield-fertility model.(DOCX)Click here for additional data file.Data S1The raw data of the crop yields of maize, barley, and soybean from the long-term fertilizer experiment from 1980 to 2010. As for the detail of the experiment, see text.(XLSX)Click here for additional data file.