In this study we applied the extreme groups/selective genotyping approach for identifying copy number variations in high and low fertility breeding boars. The fertility indicator was the calculated Direct Boar Effect on litter size (DBE) that was obtained as a by-product of the national genetic evaluation for litter size (BLUP). The two groups of animals had DBE values at the upper (high fertility) and lower (low fertility) end of the distribution from a population of more than 38,000 boars. Animals from these two diverse phenotypes were genotyped with the Porcine SNP60K chip and compared by several approaches in order to prove the feasibility of our CNV analysis and to identify putative markers of fertility.We have identified 35 CNVRs covering 36.5 Mb or ~1.3% of the porcine genome. Among these 35 CNVRs, 14 were specific to the high fertility group, while 19 CNVRs were specific to the low fertility group which overlap with 137 QTLs of various reproductive traits. The identified 35 CNVRs encompassed 50 genes, among them 40 were specific to the low fertility group, seven to the high fertility group, while three were found in regions that were present in both groups but with opposite gain/loss status. A functional analysis of several databases revealed that the genes found in CNVRs from the low fertility group have been significantly enriched in members of the innate immune system, Toll-like receptor and RIG-I-like receptor signaling and fatty acid oxidation pathways.We have demonstrated that our analysis pipeline could identify putative CNV markers of fertility, especially in case of low fertility boars.The online version of this article (doi:10.1186/s12864-015-1473-9) contains supplementary material, which is available to authorized users. Pork is the most consumed meat in the world , thus hiRecently, this level of genome variability has also been investigated in domestic animals including cattle, pig, goat, sheep, horse, dog and chicken  and holdThe applicability of SNP genotyping arrays for the estimation of DNA copy numbers have made the Porcine SNP60k chip  the methMost of the available porcine CNV studies contain functional annotations of the gene content of identified CNVRs and provide important descriptions of new individual or breed specific variants with slightly different estimates of this level of genome variation in pig populations. Furthermore, Chen et al.  has assoThe goal of this study is to investigate the feasibility of identifying candidate CNVs related to fertility in a selected population of high and low fertility boars. Gene content and reproduction QTLs that are mapped to the positions of identified CNVs were analyzed.Prior to CNV analysis several quality control steps were carried out. We have not identified any samples with outlier noise in the log R ratio values. We also checked another type of noise, the so called ’genomic waves’, that are variations of the signal intensity related to the genomic position of the probe, thus the composition of the DNA , and fouWe chose to apply the two available segmentation options in the SVS software to explore putative CNVs. This algorithm - although widely used in human studies  - has noThe first segmentation method (Univariate CNAM) searches individual genomes. Segments with significantly different log R ratio from its neighbors are identified as CNVs, which were then sorted according to the fertility status. Figure The second multivariate CNAM segmentation does not scan individual genomes, but rather checks if the segment cut-point is present in all samples for successful CNVR calling. We have used this approach to identify CNVRs specific for each fertility status by grouping the samples into low fertility or high fertility group and a 3rd control group contained all samples. As it is represented in Figure All together the two different strategies of CNAM resulted in the identification of 35 CNVRs. Fourteen CNVR were specific to the high fertility and 19 to low fertility boars  and the largest region involved in them ~12 Mb), while 5 other chromosomes had only 1–1 CNVR  was used to validate the identification of CNVRs. The results for all eight tested loci were in agreement with our predictions Figure . Every aAnother aspect of validating the predicted CNVRs was to investigate an independent set of high and low fertile animals whether the same regions could be identified. In fact, 26 CNVRs were present. Seven of them have maintained significant association with the fertility status . Interestingly, we found two regions  where samples with both gain and loss status were present, although the original predictor set of animals represented only one of them.The genomic positions of the 35 identified CNVRs were used to search for reproduction QTLs mapped to the same positions in the Animal QTLdb including the endocrine, litter size, reproductive organ and reproductive trait categories. The majority of CNVRs (30) overlapped with 137 QTLs from 16 traits and only five CNVRs are situated in regions of the porcine genome that have no reproduction QTL mapped  were also found to position within CNVRs: miR-21, miR-142, miR-143, miR-145, miR-202  are key players in gene expression regulatory networks, as such, they might be strong candidates for disease-causing non-coding sequences. The variable dosage of miRNA genes due to their involvement in CNVs is affecting their expression profile and regulatory role . Wu et amiR-202 was identified as copy number gain in low fertility and deletion in the high fertility group, which would imply a negative role in fertility regulation. This is in agreement with the observations of its marked upregulation in various testicular hystopathologic conditions  and alsoSimilar to miR-202, we have found one gene with gain/deletion copy number status in CNVR1HL. However, it is a deletion in the low fertility and a gain in the high fertility group. Although this status distribution would make it an optimal marker for fertility, it was only found in 1 animal from each group. Furthemore one gene, the Gluthatione S-transferase mu2 , is mapped to this region of chromosome 1. The superfamily of these metabolic enzymes functions as important players in protecting the cells from oxidative damage and endogenous toxicity . InteresWe have demonstrated that our analysis pipeline could identify putative CNV markers of fertility, especially in case of subfertile boars. Their relevance was demonstrated by analyzing the nature of co-localized reproductive QTLs and genes.The Canadian Centre for Swine Improvement Inc. , as a no2(Robserved/Rexpected), as described by Peiffer et al., .The data set supporting the results of this article is available in the NCBI's Gene Expression Omnibus  reposito


Color quantization is an essential technique in color image processing, which has been continuously researched. It is often used, in particular, as preprocessing for many applications. Self-Organizing Map (SOM) color quantization is one of the most effective methods. However, it is inefficient for obtaining accurate results when it performs quantization with too few colors. In this paper, we present a more effective color quantization algorithm that reduces the number of colors to a small number by using octree quantization. This generates more natural results with less difference from the original image. The proposed method is evaluated by comparing it with well-known quantization methods. The experimental results show that the proposed method is more effective than other methods when using a small number of colors to quantize the colors. Also, it takes only 71.73% of the processing time of the conventional SOM method. A variety of colors in an image provides the advantages of having greater abilities of expression and aesthetics. However, a variety of colors becomes a serious problem for most color image processing. Therefore, research to represent the various colors within a limited number of colors, which is called color quantization, has been conducted.Self-Organizing Map (SOM), which is one of the most effective color quantization methods, provides excellent results –5. HowevX denotes the original image, H and W denote image height and width, respectively.Usually, mean absolute error (MAE), mean square error (MSE), and processing time are used to quantify the performance of the color quantization results. MAE and MSE are calculated by the following, respectively:An octree is one of the image-dependent methods classified as a hierarchical clustering method. It is faster than partitional clustering methods, but its performance is relatively poor , 8.K nodes remain. The palette colors are chosen from the remaining K nodes. This method is fast and gives good results [An octree is a tree structure with up to eight nodes as children. The octree can represent all colors in an image within an eight-level tree because the colors are represented with eight bits. At first, color distribution in the image is represented using an octree, which then prunes the nodes until  results , 9.As mentioned above, octree color quantization generates the palette using the distribution of colors in the image, but it does not consider the frequency of color. This means that if an image is composed of similar colors overall but has many different low-frequency colors or noise, octree's results can be very poor.However, octree has various colors, like greenish and pinkish, which SOM does not have. This is a strength for octree for the same reason.In this paper, we present a more effective color quantization algorithm that complements the weakness of SOM by using the strengths of octree, which are fast and offer various colors.SOM color quantization shows excellent results with most images. The results are very natural and have a low MAE and MSE because the SOM learning algorithm spontaneously considers the distribution of colors.K, becomes lower), its disadvantage is apparent. The SOM learning algorithm finds the winner node and updates its weight. At that time, neighbor nodes of the winner are also updated. This means SOM generates a more natural palette, but also it means SOM loses the original color of the neighbor nodes. High-frequency colors are learned many times, so they update the weights many times. This means that high-frequency colors influence neighbor nodes much more, because when the winner is updated, the neighbor nodes are also updated. It becomes more serious when the number of colors, K, becomes smaller. If K becomes too small, then the SOM map also becomes so small that the influences on updating the winner become much greater. Therefore, the overall colors can become similar and the low-frequency colors disappear through updating the weights.However, when SOM color quantization has only a small number of colors . All of the color quantization methods were tested on an Intel i5-4460 3.2 GHz, 8.0 GB RAM machine and were implemented in C++.The proposed method was tested on a set of ten true-color images commonly used to evaluate performance. The images are shown in The color differences shown in K-means (KM) [We evaluated the proposed method by comparing it with well-known color quantization algorithms, such as popularity (POP) , octree ans (KM) , Adaptivans (KM) , and Selans (KM) .  Table K means the number of colors used for quantizing an image. The best method is indicated in bold. As expected, the experimental results show that the proposed method gives better results than the conventional SOM method when K is small. It is the consequence obtained by reducing the learning rate of the neighbors. It minimizes the influence on the neighbor by the winner, so the proposed method has the original colors of the palette. It means the proposed method is more effective than other algorithms when K is small.Tables K is large. In some cases, the proposed method gives a larger MAE and MSE, because the low learning rate of the neighbor disturbs similar color grouping on the map.On the other hand, the disadvantage of SOM is reduced when SOM uses a big enough size of map for learning, because the influence of the winner is decreased. Therefore, the performance gap between conventional SOM and the proposed method also decreases when In K is small. However, the proposed method has colors more similar to the original image because of the low learning rate of neighbors and initialization by octree colors. The proposed method gives more similar sky color on scream image and gives more similar background color on Mona Lisa image.K, is small.In this paper, an effective color quantization method is proposed. Having many colors in the original image is a serious problem for image processing, so the number of colors must be reduced. Therefore, we propose an octree-based SOM color quantization method. It is particularly more effective than other algorithms when the number of colors, K is small. The low learning rate of the neighbor means the proposed method can have colors similar to the original image.It uses an octree color quantization method to complement the disadvantages of SOM color quantization, which is one of the more effective methods. The octree color quantization method does not consider the distribution of colors, so its palette includes various colors. Therefore, we use the palette generated by octree to initialize the SOM weights. This causes the palette generated by SOM to have more varied colors than conventional SOM. In addition, the learning rate of the neighbors is set to low to minimize the loss of color information from updating the winner, because the influence of the winner becomes greater when K = 8–32). It gives lower MAE and MSE. Thus, the proposed method requires only 71.73% of the processing time of the conventional SOM method. On the other hand, when it uses a large number of colors, the performance of the proposed method gives similar MAE and MSE with conventional SOM, but it is still faster.The proposed method was evaluated by comparing it with six well-known quantization methods. MAE, MSE, and processing time are measured for comparison. The experimental results show that the proposed method is more effective than other methods when it uses a smaller number of colors to quantize the colors in the image (It is expected that the proposed method will be available to many applications that have a problem with varied colors in images.


Interferons (IFNs) activate the first lines of defense against viruses, and promote innate and adaptive immune responses to viruses. We report that the immediate early 1 (IE1) protein of human cytomegalovirus (HCMV) disrupts signaling by IFNγ. The carboxyl-terminal region of IE1 is required for this function. We found no defect in the initial events in IFNγ signaling or in nuclear accumulation of signal transducer and activator of transcription 1 (STAT1) in IE1-expressing cells. Moreover, we did not observe an association between disruption of IFNγ signaling and nuclear domain 10 (ND10) disruption. However, there is reduced binding of STAT1 homodimers to target gamma activated sequence (GAS) elements in the presence of IE1. Co-immunoprecipitation studies failed to support a direct interaction between IE1 and STAT1, although these studies revealed that the C-terminal region of IE1 was required for interaction with STAT2. Together, these results indicate that IE1 disrupts IFNγ signaling by interfering with signaling events in the nucleus through a novel mechanism.  After primary infection, human cytomegalovirus (HCMV) persists for the lifetime of the host, avoiding elimination by the host immune system . In congInterferons direct intrinsic cellular defenses and early immune responses against HCMV and other viruses. IFNs are classified into three categories: type I , type II (IFNγ) and type III (IFNl), though type I and II interferons are the best studied with respect to their antiviral functions . IFNs haThe antiviral functions of IFNs are initiated by binding with their receptors ,8,9,10. Interferons can stimulate the expression of over 300 cellular genes known as Interferon Stimulated Genes (ISGs) . The proUL123 gene forms a physical complex with STAT1 and STAT2 thereby blocking signaling after nuclear translocation and before DNA binding 32 ATP using T4 polynucleotide kinase enzyme (Invitrogen) according to the manufacturer’s recommendations. 4 µg of nuclear extract was incubated with 1× binding buffer, 0.5 µg poly dI:C, 0.1 µg poly L-lysine and 2ng of labeled probe for 15 minutes at room temperature. For competition assays, a 200-fold excess of unlabeled probe was added to the reaction mix. For supershift studies, 5 µg of rabbit polyclonal anti STAT1 antibody (Santacruz) was added to the reaction and incubated for 15 minutes at room temperature prior to adding the labeled probe. The reactions were resolved on a 4.5% native polyacrylamide gel in 0.5× TBE buffer and visualized by autoradiography.MRC5 cells were plated at a density of 1 × 10et al. [et al. reported a defect in CIITA induction by IFNγ that occurs downstream of STAT1 nuclear translocation in infected cells [+ T cells play an important role in controlling the spread of the virus. During this time it may be advantageous for the virus to be able to dampen signaling induced by IFNγ, which is produced abundantly by CD8+ T cells. This could not only counteract IFNγ–mediated antiviral responses, but also interfere with IFNγ-induced expression of MHC class I and MHC class II molecules, assembly of the immunoproteasome [The major findings of this report are as follows: (i) IE1, the immediate early transcriptional transactivator of HCMV, was identified as a candidate protein that disrupts IFN signaling by screening a HCMV cDNA library using the 2C4 reporter cell line and primary human fibroblasts; (ii) Using a series of truncation mutants we assigned this function of IE1 to the C-terminal 147 residues of the IE1 protein; (iii) IE1-mediated disruption of IFNγ signaling appears to be independent of IE1-mediated ND10 disruption; (iv) IE1 does not inhibit the proximal events of type II IFN signaling; and (v) Although IE1 does not appear to directly interact with STAT1, expression of IE1 interferes with STAT1 binding to target DNA elements. Together our data indicate that a novel mechanism is employed by IE1 to interfere with the IFNγ signal transduction pathway. In earlier studies, Miller et al.  have reped cells . Our resoteasome  and the


H2‐cells, and is therefore important in allergic disease pathogenesis. However, DCs themselves make significant amounts of TSLP in response to microbial products, but little is known about the key downstream signals that induce and modulate this TSLP secretion from human DCs. We show that human monocyte derived DC (mDC) secretion of TSLP in response to Candida albicans and β‐glucans requires dectin‐1, Syk, NF‐κB, and p38 MAPK signaling. In addition, TSLP production by mDCs is greatly enhanced by IL‐1β, but not TNF‐α, in contrast to epithelial cells. Furthermore, TSLP secretion is significantly increased by signals emanating from the endoplasmic reticulum (ER) stress response, specifically the unfolded protein response sensors, inositol‐requiring transmembrane kinase/endonuclease 1 and protein kinase R‐like ER kinase, which are activated by dectin‐1 stimulation. Thus, TSLP production by mDCs requires the integration of signals from dectin‐1, the IL‐1 receptor, and ER stress signaling pathways. Autocrine TSLP production is likely to play a role in mDC‐controlled immune responses at sites removed from epithelial cell production of the cytokine, such as lymphoid tissue.Thymic stromal lymphopoietin (TSLP) produced by epithelial cells acts on dendritic cells (DCs) to drive differentiation of T H2 immunity Thymic stromal lymphopoietin (TSLP) is a four‐helix bundle cytokine closely related to IL‐7, and was initially described as a lymphocyte growth factor H2‐cell differentiation + T cells TSLP potently activates dendritic cells (DCs), priming them to drive TAmong PRRs, dectin‐1 stimulation can induce TSLP mRNA expression in mDCs We have shown that synergistic signaling from PRR and endoplasmic reticulum (ER) stress greatly enhanced TSLP gene transcription in mDCs, similar to the effect of ER stress on IL‐23p19 expression The ER senses dangers to internal homeostasis and initiates the unfolded protein response (UPR) when the protein folding capacity of the cell is exceeded causing disruptions to normal protein folding mechanisms. The UPR is controlled by the three principal ER stress sensing proteins, inositol‐requiring transmembrane kinase/endonuclease 1 (IRE1α), protein kinase R‐like ER kinase (PERK), and activating transcription factor 6 Here, we show that dectin‐1 signaling stimulates TSLP production from mDCs and that this is enhanced by signals from the ER stress sensing proteins, IRE1α, and PERK. In addition dectin‐1‐induced IL‐1β is required for optimal TSLP secretion; however, we show that dectin‐1‐induced TSLP requires independent, mutually exclusive signals emanating from the ER stress response and the IL‐1 receptor.Alcaligenes faecalis), smaller particulate β‐1,3 glucan microparticles (MPs) derived from Saccharomyces cerevisiae, and the hyphal yeast, Candida albicans. Dectin‐1 signaling stimulated via C. albicans or β‐glucan particle engagement induced TSLP secretion in immature mDCs . Our findings showed that dectin‐1‐induced inflammasome/caspase‐dependent IL‐1β is required for optimal TSLP production from mDCs; however, inflammasome‐independent signaling events are also required.Processing and secretion of IL‐1β requires inflammasome activation mediated by caspase recruitment and activation SLP Fig. E, and ILSLP Fig. F. FurtheAlthough Syk, NF‐κB, and p38 MAPK signaling is required for IL‐1β expression, the addition of exogenous IL‐1β could not rescue TSLP expression in the presence of these inhibitors . Therefore, the reduction of TSLP expression in the presence of these inhibitors cannot be solely attributed to the loss of IL‐1β.We have previously presented microarray data showing that ER stress can augment PRR‐induced TSLP and IL‐23p19 in mDCs + DC  or thapsigargin in conjunction with dectin‐1 stimulation increased TSLP secretion from mDCs Fig. A, and fr DC Fig. B. Import DC Fig. A, showinER stress results in the activation of three key sentinel proteins, IRE1α, PERK, and activating transcription factor 6 The ISR can also be activated by other eIF2α kinases including protein kinase R (PKR). Although we show that β‐glucan stimulation potently activated PKR , inhibition of PKR had no effect on β‐glucan‐induced TSLP . This suggests that activation of PERK is critical for TSLP induction.The ability of IRE1α or PERK inhibitors to reduce TSLP secretion suggests that ER stress signals were regulating TSLP expression at the level of gene transcription. Both IRE1α and PERK inhibition reduced TSLP mRNA expression during dectin‐1 stimulation without altering its kinetics of expression Fig. F. In conThe ability of ER stress to enhance TSLP expression raises the question whether IRE1α‐ and PERK‐regulated signals exhibit their effect indirectly by modulation of IL‐1β expression. Inhibition of IRE1α but not PERK activation reduced IL‐1β secretion from mDCs Fig. A. The reTranscription factors induced during the UPR may be involved in the induction of TSLP gene expression. We have previously shown that ISR‐induced C/EBP homologous protein (CHOP) is a critical transcription factor in IL‐23p19 expression C. albicans or β‐glucans is regulated by an integration of signals from dectin‐1, the ER stress response, and the IL‐1 receptor.DCs are professional antigen‐presenting cells and have the ability to sense infection and danger signals and instruct immune cells to differentiate into different cell subsets. TSLP is an important cytokine in immune regulation 5 cells). It is difficult to directly compare TSLP production by mDCs and epithelial cells/fibroblasts; however, stimulation of these cell types typically generates ∼50–300 pg/mL from ∼2–4 × 105 cells + DC upregulate TSLP mRNA to a higher level compared to epithelial cells when both cell types are challenged with a stimulus to which both can respond C. albicans and β‐glucan, though they may produce lower amounts than their human counterparts. It is likely that TSLP secretion is highly stimulation specific. Therefore, mDCs should not be discounted as potentially important sources of TSLP and we suggest this TSLP could have an important function during an initial fungal challenge.We noted that mDCs generated significant levels of TSLP/cell  quantities of this cytokine H2/regulatory T cells or modulate the differentiation of TH17 cells We have shown clear differences in the quantities of IL‐1β induced by different dectin‐1 agonists, whereas TSLP production remained stable. Pioneering experiments that established TSLP as a critical TC. albicans or β‐glucans produce TSLP and that this process involves the integration of signals from dectin‐1, the ER stress associated proteins IRE1α and PERK, and the IL‐1 receptor. These data argue that these signaling pathways may provide novel therapeutic targets to promote or downregulate TSLP production in relevant clinical settings.In summary, we have demonstrated that dectin‐1 stimulation of mDCs by + monocytes were purified from human PBMC by magnetic bead separation (Miltenyi) and were differentiated into mDCs with 20 ng/mL GM‐CSF (Life Technologies) and 4 ng/mL IL‐4 (BD Biosciences) by culturing for 6 days in RPMI1640 5% FCS as described previously + DC were isolated from human PBMC by magnetic bead separation (Miltenyi). Murine BMDCs were isolated from the femurs of wild‐type (WT) and CHOP KO C57B6 mice and cultured for 7 days in RPMI1640 10% FCS supplemented with 5% X63 conditioned media and 10 ng/mL IL‐4 (Peprotech).CD145/mL and were stimulated with 50 μg/mL of the β‐glucans, curdlan derived from A. faecalis , β‐1,3 glucan MPs derived from S. cerevisiae  or heat‐killed hyphal C. albicans  grown in sabouraud dextrose broth for 8 h at 37°C with shaking to an optical density of 0.2. Candida albicans was killed by heating for 1 h at 70°C.mDCs were seeded at a density of 2.5 × 102B dectin‐1‐blocking antibody (clone‐259931 R&D Systems), 10 μg/mL mouse IgG2B isotype control (clone‐20116 R&D Systems), 5 μM Syk inhibitor, R406 (Selleckchem), 10 μM NF‐κB inhibitor, isohelenin , 1 μM p38 MAPK inhibitor, BIRB0796 (Selleckchem), 5 μg/mL anti‐TNF‐α, Infliximab (Jannsen Biotech), 1 μg/mL IL‐1 receptor antagonist , 50 μM caspase‐1 inhibitor (Z‐YVAD‐FMK) , 50 μM caspase‐8 inhibitor (Z‐IE(OMe)TD(OMe)‐FMK) , 10 ng/mL IL‐1β (Miltenyi), 1 μg/mL TM (Sigma‐Aldrich), 0.25 μM thapsigargin (Sigma‐Aldrich), 30 μM IRE1α inhibitor, 4μ8C , and 1 μM GSK‐PERK inhibitor (Toronto Research). Where inhibitors, blocking antibodies and modifiers were used, mDCs were pretreated 1 h prior to cell stimulation. Repeated experiments were performed on independent donors unless otherwise stated.Ten micrograms per millimeter mouse IgGmDC culture supernatants were harvested after 24 h of stimulation. The concentrations of TSLP (R&D Systems) and IL‐1β (eBioscience) were measured by ELISA as per the manufacturer's instructions.Total RNA was extracted from cells using RNA Mini Kits (Bioline). Quantitative real‐time PCR was carried out using TaqMan Gene Expression Assays (Applied Biosystems) for TSLP, IL‐1β, and OPG and the SYBR Green method for XBP1 splicing. TSLP (Hs00263639_m1), IL‐1β (Hs00174097_m1), and OPG (Hs00171068_m1) TaqMan PCR probe primer sets (Applied Biosystems), and HPRT (MWG‐Biotech), probe, 5′‐TTCCTCATGGACTAATTATGGACAGG‐3′, forward, 5′‐TGAGGATTTGGAAAGGGTGTTT‐3′ reverse, 5′‐ACATCTCGAGCAAGACGTTCAG‐3′. SYBR primer sets (MWG‐Biotech), XBP1 spliced, forward 5′‐TGCTGAGTCCGCAGCAGGTG‐3′, reverse 5′‐GCTGGCAGGCTCTGGGGAAG‐3′, HGPRT, forward 5′‐GACACTGGCAAAACAATG‐3′, reverse 5′‐ACAAAGTCTGGCTTATATCC‐3′. Gene expression was normalized to HPRT and calculated as fold change from unstimulated control.mDCs cell were lysed on ice in 250 μL of ice cold cytoplasmic lysis buffer . After lysis the cytoplasmic extract was frozen at −20ºC overnight before thawing to aid cell lysis. Lysates were then centrifuged for 15 min at 4°C and the supernatant retained. Bradford assay was carried out to determine protein quantification (Thermo) and proteins were separated by SDS‐PAGE. Resolved proteins were detected after transfer by immunoblot with indicated primary antibodies, followed by incubation in HRP‐conjugated secondary antibodies and ECL (PerkinElmer) and visualized using GBox (Syngene). Monoclonal antibodies for phospho eIF2α (S51) , eIF2α , phospho PKR , PKR , IκBα , phospho Syk (Tyr525/525) , Syk , phospho p38 MAPK (Thr180 and 182) , p38 MAPK , and β‐actin  were used for detection.mDCs were resuspended in ice cold PBS supplemented with 2 mM EDTA, 0.1% BSA, and 0.01% sodium azide, blocked with 10% mouse serum (Sigma) and stained with anti‐CD83 (BD), anti‐CD86 (BD), and anti‐HLA‐DR (Biolegend). Surface expression was evaluated relative to unstimulated stained controls on a BD CANTO II and analyzed using FlowJo software (Treestar).t test  or one/two‐way ANOVAs with Bonferroni post‐tests *p < 0.05, **p < 0.01, ***p < 0.001.Data were analyzed using GraphPad Prism statistical package. Cumulative data are displayed as mean ± SEM. Statistical analysis using either The authors declare no commercial or financial conflict of interest.BMDCBM‐derived DCCHOPC/EBP homologous proteineIF2αeukaryotic initiation factor 2 αHGPRTHypoxanthine‐guanine phosphoribosyltransferaseIRE1αinositol‐requiring transmembrane kinase/endonuclease 1ISRintegrated stress responsemDCmonocyte‐derived DCMPsmicroparticlesOPGosteoprotegerinPERKprotein kinase R‐like ER kinaseTSLPthymic stromal lymphopoietinUPRunfolded protein responseXBP1X‐box binding protein 1As a service to our authors and readers, this journal provides supporting information supplied by the authors. Such materials are peer reviewed and may be re‐organized for online delivery, but are not copy‐edited or typeset. Technical support issues arising from supporting information (other than missing files) should be addressed to the authors.Peer review correspondenceClick here for additional data file.Supporting FiguresClick here for additional data file.


The idea that trade-offs between offspring quantity and quality shape reproductive behaviour has long been central to economic perspectives on fertility. It also has a parallel and richer theoretical foundation in evolutionary ecology. We review the application of the quantity–quality trade-off concept to human reproduction, emphasizing distinctions between clutch size and lifetime fertility, and the wider set of forces contributing to fertility variation in iteroparous and sexually reproducing species like our own. We then argue that in settings approximating human evolutionary history, several factors limit costly sibling competition. Consequently, while the optimization of quantity–quality trade-offs undoubtedly shaped the evolution of human physiology setting the upper limits of reproduction, we argue it plays a modest role in accounting for socio-ecological and individual variation in fertility. Only upon entering the demographic transition can fertility limitation be clearly interpreted as strategically orientated to advancing offspring quality via increased parental investment per child, with low fertility increasing descendant socio-economic success, although not reproductive success. We conclude that existing economic and evolutionary literature has often overemphasized the centrality of quantity–quality trade-offs to human fertility variation and advocate for the development of more holistic frameworks encompassing alternative life-history trade-offs and the evolved mechanisms guiding their resolution. In the conventional demographic literature, the concept is typically accredited to the economist Gary Becker. Before Becker, fertility was widely considered to be outside the realm of economic analysis . In partet al.  demonstra priori in economics in evolutionary models of organic diversity [Less well known to many demographers, the quantity–quality trade-off concept has a parallel and theoretically richer origin in evolutionary ecology. Evolutionary life-history theory accounts for both species and individual-level variation in reproductive strategies in terms of the allocation of resources  to competing functions that define the life cycle of an organism –11. Sinciversity . It alsoiversity . Economiiversity . Many deiversity ,12,15).within the potential phenotypic range of fertility for our species, the optimization of resource allocation to offspring quantity versus quality can account for observed socio-ecological and individual-level variation in fertility.The objectives of this review are to: (i) overview the origin and current application of the quantity–quality trade-off concept in evolutionary ecology; and (ii) consider the strengths and limitations of this concept in accounting for human fertility variation in both pre- and post-demographic transition populations. Humans fall on the extreme end of already high mammalian parental investment and have highly altricial offspring. It thus seems intuitive that trade-offs between offspring quantity and quality will be fundamental to human life-history evolution –18. ThroWe begin by summarizing the contributions of David Lack, a pioneer of life-history theory, to the study of clutch size variation in birds, and the legacy of his research in contemporary studies of animal and human reproductive behaviour. From this platform, we argue that Lack's original focus on clutch size rather than lifetime fertility, in addition to economic demography's focus on the demographic transition rather than variation in pre-demographic transition fertility, has led much of the existing literature to overemphasize the pivotal role of the offspring quantity–quality trade-off in accounting for human fertility variation. This position is supported from a review of the anthropological literature concerning high fertility and high mortality populations somewhat characteristic of our evolutionary past. Here, we argue that a number of important factors reduce costly sibling competition across the life course, so that the relevance of quantity–quality trade-offs to the initiation, continuation, or termination of reproduction is likely to be relatively modest compared with populations that are undergoing or have completed the demographic transition. We then briefly discuss alternative life-history trade-offs that may play a more dominant role in determining the individual variance of fertility in pre-demographic transition environments. To conclude, we revisit the demographic transition and consider how a more holistic appreciation of the multiple life-history trade-offs and selective forces that influence fertility identifies gaps in our current understanding and opportunities for future research.2.(a)species [individual level, so that individuals reproducing at a lower rate than that which maximizes reproductive success are rapidly outcompeted [Clutch size has been evolved through natural selection to correspond with the largest number of young for which the parents can on the average find enough food. , p. 32This simple hypothesis was one of the first optimization models applied to the study of animal behaviour, contributing substantially to the transformation of ecology from a descriptive discipline to a predictive and experimental science, and earning Lack the epithet ‘father of evolutionary ecology’ [For evolutionary demographers, the idea that contingent trade-offs between offspring quantity and quality determine individual variation in fertility has its roots in the work of the ornithologist David Lack. Lack was among the very first researchers to initiate the application of the ‘Modern Synthesis’, the successful integration of Darwin's theory of natural selection with Mendelian genetics, to the study of animal behaviour . Before species  .competed . In his ecology’ .Lack tested his ideas using a novel experimental method, the manipulation of clutch size by transferring newly hatched young to and from nests to create enlarged and reduced broods. Clutch size manipulations have now been conducted on a wide range of avian species where robbing and/or cuckolding parents is a relatively straightforward procedure . The met(b)First, Lack was concerned with the optimization of a single reproductive bout. However, in iteroparous species , investment in one clutch will also influence available resources for future survival and reproduction. In other words, considering a single clutch ignores trade-offs between reproduction and somatic maintenance, and consequently between current and future reproduction . This haet al. [Second, Lack's account implicitly assumes asexual reproduction and uniparental care by neglecting how the dynamics of partner attraction, retention, and coordinated offspring provisioning may influence the resources available for investing in offspring quantity and quality, and the costs and benefits thereof (e.g. ). Parentet al.  and the et al. on ‘evolutionary traps’ [Third, Lack's evolutionary ecological model identified the optimal clutch size as that of a population assumed to be in equilibrium. This reliance on the assumption that appropriate mechanisms exist which adapt behaviour to the current environment may be unwarranted. Contemporary evolutionary approaches to behaviour incorporate a consideration of the necessarily imperfect mechanisms making up an organism's ‘adaptive toolbox’, providing predictions about when and why we should anticipate departures from strict optimality . Two poiy traps’ ) it is ay traps’ , and it y traps’ .Finally, both life-history theory and economic accounts of fertility, along with related sociological models of family size and achievement ,42, are 3.Just as studies of clutch size were pioneering for evolutionary studies of animal behaviour, some of the first anthropologists to apply evolutionary models of behaviour, i.e. human behavioural/evolutionary ecologists, focused on variation in fertility . Below, (a)Available evidence indicates high fertility often compromises child survival in pre-demographic transition settings, but rarely to the extent that reducing fertility can be understood as a strategy to maximize individual lifetime reproductive success . Child survival is under strong selection in human life-history evolution . In highIn part motivated by this surprising result, a spate of later studies, mostly concerning contemporary small-scale agriculturalist or historical agrarian populations, went on to examine relationships between fertility and child survival. These studies do show that high fertility is associated with low child survival –68. HoweOne reason why trade-offs between fertility and child survival may be seemingly absent, or at least insufficiently strong to favour fertility limitation, is the problem of phenotypic correlations . Accordiet al. [There are additional reasons why trade-offs between fertility and child survival may be weak, so that parents do not optimize their fertility decisions on the basis of expected mortality. Child mortality may be high but ‘extrinsic’, i.e. largely care-independent within locally feasible ranges of parental investment, as opposed to ‘intrinsic’, i.e. largely tied to variation in parental investment . This doet al. , in an a(b)Evidence that high fertility diminishes offspring capital is mixed in pre-demographic transition populations, with many studies indicating that increasing fertility can be beneficial at even very high levels. With regard to ‘embodied capital’ , some studies demonstrate that children in larger families have relatively poor physical health, but others find no trade-off or mixed effects depending on sibling characteristics and/or health measures used –77. DataA number of factors may limit costly sibling competition over embodied capital in developing and ‘traditional’ populations. First, while there is probably always a net energetic deficit to rearing children , childreAvailable evidence suggests ‘relational capital’  is largely not subject to dilution effects with increasing sibship size. Instead, larger numbers of siblings appear to increase an individual's network of altruists who may provide support across multiple dimensions. Draper & Hames , for exa(c)Sibling competition between surviving offspring is only predicted to lead to fertility limitation if it ultimately diminishes offspring reproductive success. To date, trade-offs between offspring number and offspring reproductive success among survivors have only been demonstrated in contexts where material wealth is transmitted across generations, i.e. stemming from the domestication of plants and animals . For exaWhere substantial material wealth is inherited , and where sibling competition potentially reduces reproductive success, formal mathematical models have indicated that the costs of raising and marrying off children could hypothetically favour fertility limitation to maximize fitness ,89. Howe4.In the sections above, we have made the argument that, with the exception of avoiding substantial costs of tight birth spacing on infant survival, the costs of high fertility on offspring quality appear largely offset by the benefits of greater offspring quantity, at least in the observed range of our species . In this(a)In the absence of intense competition between offspring for post-natal investment, fertility in pre-demographic transition societies may be best understood as primarily determined by the availability of energetic reserves required for offspring production in pregnancy—redirecting our attention to the life-history trade-off between reproductive and somatic effort. This is particularly true for income breeders  where inet al.'s analysis [Furthermore, age at first birth in humans reflects a trade-off between the costs of delay (truncating the remaining years available for reproduction), which are elevated in high-mortality environments, with the benefits of postponement via the accruement of personal capital (such as enhanced education and income) . The cleanalysis  .within pre-demographic transition contexts, where fertility variation is substantial but still poorly understood from an evolutionary perspective [Positioning the trade-off between reproduction and somatic effort as pivotal to reproductive decision-making suggests resource access and maternal energy budgets are fundamental determinants of socio-ecological variation in fertility, rather than the extent to which local ecologies dictate high or low levels of costly sibling competition. As noted above, maternal energy budgets are likely to be particularly pertinent in a species where foraging and/or food acquisition and preparation is a daily concern and can be energetically expensive. A simple prediction might be that better socio-ecological conditions would lead to higher fertility—since more resources are now available for reproduction. However, the situation is complex because better conditions may not only improve maternal energy budgets, but also increase the survival chances of children, thereby increasing the returns to higher investment and altering patterns of replacement and insurance fertility. As we describe below, the demographic transition also clearly bucks the expectation that resource-rich ecologies will lead to greater fertility. More research is required to assess between population variation in human fertility spective ,113 and (b)Despite the common simplifying assumptions of many formal models of fertility in both economics and evolutionary anthropology, human reproduction is a sexual act, thereby entailing the fitness interests of more than one individual. Measures of life-history trade-offs should accordingly incorporate currencies earned through sexual selection. This will produce a more fully unified evolutionary account of fertility that includes social and institutional considerations and constraints ,115. ForIn a species characterized by relatively stable pair bonds, individuals face a trade-off between starting to reproduce early and waiting to find a preferred mate. Accordingly, variation in age at marriage probably reflects the dynamics involved in mating effort, sensitive to opportunities to get a high-quality mate who can provide both direct and indirect fitness benefits. Such concerns will be particularly relevant when reproduction involves significant resource transfers within the pair bond, again characteristic of human marriage, where males and females typically engage in relatively complementary economic activities . For exaTurning from first marriage to mate switching, it may pay to slow down reproduction if searching for a replacement mate, and to enhance fertility if aiming to retain a current mate, at least from female point of view. This is because typically remarriage is more difficult for women with children  and divoet al. [While the trade-off between reproduction versus some aspects of maternal somatic capital is mainly regulated by physiology, social institutions can play a larger role in determining how mating dynamics influence fertility. For example, if men have the option of polygyny, there may be less pressure on women to produce offspring at a faster rate than is their preference . Similaret al.  propose 5.As populations undergo socio-economic and cultural ‘modernization’, the factors that once reduced costly sibling competition begin to erode. Children are no longer involved in active resource production and so have little potential to underwrite their own costs to parents. Kin networks become fragmented, and the emergence of low fertility rates leads to a lower absolute number of potential alloparents . Decliniperception of quality. This suggests there is adaptive lag in mechanisms governing fertility. Under modernization, humans elevate their perception of the costs of high fertility on offspring capital, leading to a correspondingly exaggerated strategy of fertility limitation [et al. propose that that:Because human parents and grandparents provision children, natural selection probably produced mechanisms by which fertility could respond to the number of children parents could afford to raise in any given socio-ecology. , p. 238In modern environments such adaptive mechanisms then lead to maladaptively low fertility in response to increased perceived costs of child rearing well outside the range experienced previously by our ancestors.Many evolutionary demographers have argued that modern fertility decline may be adaptive as part of an optimal regulation of the quantity–quality trade-off, provided substantial economic rewards are bestowed on descendants ,131,132.mitation ,35,136. On the basis of the literature reviewed across previous sections, we highlight two important considerations stemming from a more holistic appreciation of the multiple selective forces acting on human-fertility variation. First, if it is indeed true that modern fertility limitation is best understood as a strategy to advance offspring status in line with a perceived quantity–quality trade-off, then this represents a potentially radical shift. As we have argued, there is little indication that fertility variation can be accounted for by the tactical balancing of equivalent forms of this trade-off in pre-demographic transition environments. In this sense, limiting fertility to strategically enhance offspring success cannot be considered a straightforward extension of pre-existing reproductive strategies. This raises very interesting questions about the flexibility of human behaviour under rapidly changing environments, and identifies the need for a more developed study of the cognitive mechanisms underlying human reproduction to match the substantial progress made in our understanding of the physiology of human reproduction across the 1980s and 1990s . Clearlyet al. [Second, we caution that while data on the apparent benefits of low fertility to descendant material capital certainty appear consistent with the view that perceived quantity–quality trade-offs drive the demographic transition, evolutionary anthropologists' persistent preoccupation with this trade-off means that relatively little attention has been paid to the possibility that alternative trade-offs play a substantial role in modern fertility patterns. Indeed, it remains an open possibility that modern low fertility may be better understood in terms of exaggerated returns of low fertility to own socio-economic success relatively independently of the socio-economic consequences of costly sibling competition. Accordingly, we should perhaps refocus our attention more on the factors that lead individuals to postpone fertility, i.e. the decision to reproduce now versus later, rather than the decision of how many offspring to have . This peet al.  review, 6.The optimization of the life-history trade-off between offspring quantity and quality is surely fundamental in defining the theoretical upper limits of human fertility and our propensity for singleton births. Yet the extent to which it can meaningfully account for why so few women approach maximal fertility and for the substantial ecological and individual variance in fertility rates observed even before the demographic transition remains an open question. Our review of sibling competition in high-fertility, high-mortality populations leads us to propose that alternative life-history trade-offs, such as that between reproductive and somatic effort, and between fertility and mating effort, play a more pivotal role in accounting for fertility variation in settings characteristic of our evolutionary past. Even where high fertility has notable costs on offspring reproductive success due to substantial material wealth transfers at marriage and inheritance, a relatively novel form of sibling competition following the domestication of plants and animals, this dilemma has typically been solved by biased parental investment rather than tactical fertility reduction.The trade-off between the perceived quality and quantity of offspring is more obviously relevant to decisions to reduce fertility accompanying socio-economic and cultural modernization, an observation that may account for our preoccupation with this trade-off, since researchers themselves are living in environments where the direct and opportunity costs of raising children are particularly salient. However, even in modern low-fertility contexts, we suggest that expanding present theoretical frameworks beyond the persistent yet artificial single clutch and asexual reproduction assumptions, dating back to both Lack and Becker, will be necessary to account for observed fertility patterns. The quantity–quality trade-off concept has propelled a vast literature addressing species, population, and individual-level variation in reproductive strategies. Nevertheless, we will ultimately understand a lot more about reproductive decision-making by enriching current frameworks with greater attention to weighing up the relative contribution of the quantity–quality trade-off to the alternative selective forces discussed in this paper.


The overall decline of fertility in Amhara National Regional State between 2000 and 2011 was the highest in Ethiopia. The aim of the present study was to determine the most significant proximate determinant of fertility change during the last decade in the region using Bongaarts’ model.The sources of data were the 2000, 2005, and 2011 Ethiopia Demographic and Health Surveys. The model indices were calculated for each survey. Decomposition of fertility change into components of proximal determinants was also carried out. An index value close to 1 is a negligible inhibiting effect while a large inhibiting effect when the value very closes to 0.The fertility-constraining effect of contraception increased from 0.93 in 2000 to 0.65 in 2011; however, it was lower than the effect of postpartum insusceptibility at all given times. The index of marriage remained unchanged in constraining fertility over the period (0.71 in 2000 and 0.70 in 2011) while the influence of postpartum insusceptibility slightly declined from 0.49 in 2000 to 0.54 in 2011 but was stronger than contraception and marriage. The contribution of contraception was most important in urban areas (0.46 in 2011 from 0.52 in 2005 and 0.64 in 2000); however, in rural areas, it became an important determinant over the period (0.95 in 2000 and 0.69 in 2011). The effect of postpartum insusceptibility in rural areas showed a decreasing trend (0.48 in 2000 and 0.53 in 2011). The index of marriage in rural areas was stable overtime (0.75 in 2000 and 0.73 in 2011) while in urban areas the effect declined from 0.42 in 2000 to 0.65 in 2011. Marriage was the most important proximate determinant of fertility among women with secondary and above education but the impact declined during the period (0.41 in 2000 and 0.61 in 2011). The importance of postpartum insusceptibility in limiting fertility among women with secondary and above education declined overtime (0.77 in 2000 and 0.87 in 2011) whereas the contribution of contraception became more important (0.44 in 2000 and 0.35 in 2011).An increase in the level of contraceptive use and effectiveness overtime was the single most important contributing factor for the recent fertility decline in the region. Fertility levels and trends in the world generally and in a country particularly vary across time. The casual factors are diverse, but they may be broadly grouped into direct or proximate determinants and indirect or background factors . The proFurther, Bongaarts’ model has quantified the contribution of these four factors to the observed fertility level. This resulted in a simple but powerful model for analyzing how fertility changes overtime or differs from one group to another; therefore, it is the most extensively used models in the world to measure the extent to factors reduce fertility. The proximate determinants model is mostly used to identify the main factors behind fertility changes, the changes associated with fertility through time, and to compare changes in fertility among countries or regions of a country .The National Population Policy of Ethiopia was adopted in 1993 with an overall goal of harmonizing the rate of population growth and economic development primarily addressing the high fertility. The policy in particular, aims to achieve a total fertility rate of 4.0 children per woman in 2015. The policy envisages meeting this target by employing different strategies such as expanding clinical and community-based contraceptive distribution services; promoting breastfeeding as a means of increasing the time span between pregnancies; and raising the minimum age at marriage for girls from the current lower age limit of 15 to, at least, 18 years  and the With a population of nearly 98.1 million in 2015, Ethiopia is the second most populous country in Africa next to Nigeria and is projected to be 165.1 million in 2050 . The popFertility declined in all regions of the country except Afar, Somali, and Benishangul-Gumuz National Regional States during the last decade. It remained constant in Afar National Regional State (4.9 vs. 5.0 children per woman) and Benishangul-Gumuz National Regional State (5.4 vs. 5.2 children per woman) between 2000 and 2011. However, in Somali National Regional State the total fertility rate increased from 5.7 children per woman in 2000 to 7.1 children per woman in 2011 –16. In AThe available studies on proximate determinants of fertility in Ethiopia were carried out at a national level of aggregation , 18; theDetailed study on the trends in proximate determinants of fertility change is important for assessing the implementation of population/reproductive health policies and family planning programs and the efforts achieving the overall goal of changing fertility behaviour, that is, increasing contraception, reducing unmet need for family planning, and rising age at first marriage. In addition, the decomposition of proximate determinants is also a basis for future projections of changes in the overall level of fertility which may be expected as a result of changing socioeconomic, demographic, cultural, institutional, psychological, health, and environmental conditions.The 2000, 2005, and 2011 Ethiopia Demographic and Health Surveys were the data sources for this analysis. They were designed to provide population and health indicators at the national and sub-national levels. The sample was selected using a stratified two-stage cluster design. Enumeration areas were the sampling units for the first stage and households comprised the second stage of sampling. A complete listing of households was carried out in each of the selected enumeration areas and a representative sample of households was selected proportionally. All women of reproductive age found in the selected households were interviewed. A representative sample of 2059, 2158, and 2163 households were selected for the 2000, 2005, and 2011 survey, respectively, in Amhara National Regional State. The 2000, 2005, and 2011 Ethiopia Demographic and Health Surveys successfully interviewed 3820, 3482, and 4433 women of reproductive age, respectively, in the region.Bongaarts proposed the proximate determinants of fertility model at a time when it was assumed there was very little intercourse outside of marriage. Hence, all of the calculations he proposed were based only on currently married women of reproductive age. Bongaarts initially proposed the proximate determinants framework; social and cultural mores have been shifting to greater or lesser degrees across the world and girls and women are now having sexual intercourse before and outside marriage. Stover proposed updating the framework to account for the fact women are having sex before and outside marriage and these same women may use contraceptives and seek induced abortions . This stA)Role of proximate determinants of fertilityIn addition, when contraceptive use is concentrated at later ages, the Bongaarts’ model is a poor estimator of the fertility-inhibiting effect of contraception because women use contraception to stop rather than to space births . These pThe model indices were calculated for each survey separately. The model expresses the total fertility rate as a product of four indices  and total fecundity rate , 3, 20. TFR is the total fertility rate, TF is the total fecundity, Cm is the index of marriage, Cc is the index of contraception, Ca is the index of induced abortion, and Ci is the index of postpartum insusceptibility.Where, Each index has a value between 0 and 1. When the index is close to 1, the proximate determinant has a negligible inhibiting effect on fertility, whereas, when it takes a value very close to 0, it has a large inhibiting effect , 3. Thati)TF)Total fecundity Cm)Index of marriage  that the total fertility rate would take if all the indices of the proximate determinants were 1. That means, a fertility level that would prevail if all women age 15–49 were married, there was no contraception use in the population concerned, no postpartum insusceptibility (beyond a maximum of 1.5 months), and no induced abortion , 3. AccoThe index of marriage represents the proportion by which total fertility rate is smaller than the total marital fertility rate as a result of marital pattern (the effect of non-marriage in terms of a reduction in fertility per women). The higher the level of marriage in the population, the less the inhibiting effect on fertility and vice versa. The index of marriage is calculated as the proportion of total fertility rate over the sum of age-specific marital fertility for a given population at a given time, that is,Cm is the index of marriage, m(a) is the age-specific proportions of women currently married and is produced by dividing the number of married women of a particular age group by the number of women in the same age group and g(a) is the age-specific marital fertility rate and computed by dividing the age-specific fertility rate by the proportion of women currently married in each age group.iii)Cc)Index of contraception . In terms of the marital fertility rate, the index of contraception gives the proportion by which the total marital fertility rate is smaller than total natural fertility (fertility in the absence of any deliberate birth control practice). However, the contraception index value depends on the current contraceptive prevalence rate and the average use effectiveness of the contraception. The index of contraception varies inversely with the contraception prevalence and use effectiveness of modern contraception practiced by couples. The index of contraception is calculated as the proportion of currently married women of reproductive age currently using specific methods of modern contraception, that is,Cc is the index of contraception, u is the contraceptive prevalence among married women (including those living together as married) age 15–49, and e is the average effectiveness of contraception. The constant 1.08 given in Eq. ) level, the weights being the proportion of women employing a specific method (u(i)), that is,The value of iv)Ci)Index of postpartum insusceptibility  plus the duration of postpartum insusceptibility. The constant 20 in Eq. ( + 9 plusCi is the index of postpartum insusceptibility and i is the mean duration of postpartum infecundability measured in months.B)Decomposition of Change in Proximate Determinants of FertilityWhere, The decomposition of proportional change in the total fertility rate into the components of the proximate determinants was also carried out. The purpose of decomposition analysis of proximate determinants was to identify the sources of changes in the predicted total fertility in the last decade. A decomposition method developed by Kitagawa and later elaborated by Das Gupta was used to analyze the data , 24. TheStep 1: Decomposition of change in total fertility rate over the time frame can be calculated as:f)Proportional change in total fertility rate m)Proportional change in total fertility rate due to a change in the index of marriage (Pc)i)Proportional change in total fertility rate due to a change in the index of postpartum insusceptibility (Pd)c)Proportional change in total fertility rate due to a change in the index of contraception (Pe)r)Proportional change in total fertility rate due to a change in the remaining proximate variables  - fecundability, intrauterine mortality, and sterility Table A skewed method-mix had been apparent in the region with only three methods, namely injectables, implants, and pills accounting for about 97% of all contraceptive use in 2011. Injectables were by far the most predominant method in all surveys. The share of injectables rose over the last decade, from 52% in 2000 to 75% in 2005 and 80% in 2011. Between 2000 and 2005 the proportion of implants to the total contraceptive use remained constant at 2% but there was an increase to 12% in 2011. In 2011 only 5% of total contraceptive use was the pill and it was 45% and 23%, respectively, in 2000 and 2005. Other methods taken together contributed to less than 3% of overall contraceptive use in 2011.In Amhara National Regional State, the median age at first marriage among currently married women increased slightly from 14.5 years in 2000 to 15.1 years in 2011. The proportion of women who married at age 15, 18 and 20 declined from 56.5%, 87.0%, and 92.1% in 2000 to 48.3%, 74.9%, and 83.0% in 2011, respectively. The early marriage was highest in the country though it showed only a 12% decline in the last decade from 87% in 2000.In Amhara National Regional State, the mean duration of amenorrhea declined from 22.4 months in 2000 to 18.4 months in 2011 in the total sample of currently married women. The mean duration of abstinence in the total sample of currently married women increased from 2 months in 2000 to 2.7 months in 2011. The mean duration of insusceptibility declined from 22.6 months in 2000 to 19.0 months in 2011.Table The fertility-constraining effect of contraception increased as evidenced by a decrease in the index value overtime. The estimated contraception index declined from 0.93 to 0.65 during the period. The fertility-constraining effect of postpartum infecundability decreased as evidenced by an increase in its index value overtime. The estimated index of postpartum infecundability increased from 0.49 in 2000 to 0.54 in 2011. The index of marriage overtime remained unchanged with an index value of 0.71 in 2000 and 0.70 in 2011.Table In 2000, postpartum insusceptibility and marriage were seen the primary fertility-limiting factors for illiterate women (0.48) and women with secondary and above level of education (0.41), respectively. Contraception also played a role in limiting fertility among women with secondary and above level of education (0.44) while its impact was minimal among illiterate women (0.95). A similar pattern was observed in 2005 and 2011 except the fact that contraception had gained prominence in limiting fertility among women with a level of education above primary school (0.35). Moreover, contraception showed a large shift of impact in controlling fertility among illiterate women and women with primary level of education.Once the indices of the three major proximate determinants had been calculated for each survey, the next step is to decompose the change in the total fertility rate observed between two time points to determine contributions of the three proximate determinants Table . A decliThe changes in the total fertility rate by education and residence were also decomposed by the proximate determinants. There was a consistent increase in the total fertility rate in urban areas whereas fertility in rural areas of the region had declined. The largest portion of the increase in fertility of urban areas between 2000 and 2011 happened as a result of a change in the proportion of married women of reproductive age over the period (53.4%) despite the fact that postpartum insusceptibility (−2.1%) and contraception (−28.0%) were associated with a decline in fertility. The percent change in the total fertility rate of rural women during 2005–2011 (−16.2%) was twice that observed during the first-half of the decade (−8.3%). Change in the proportion of rural women using modern contraception was responsible for a significant proportion of the decline observed in fertility (−26.7%) over the period 2000 to 2011.From 2000 to 2005, the decline in fertility observed among women with no formal education was larger than women with above primary education. The responsible factor for this decline was a change in the prevalence of contraception over the period among illiterate women, however, the decline in fertility among women with secondary and above level of education was associated with factors that were not included in the model (interaction = −66.8%). On the contrary, the increase in fertility among women with primary level of education (25.2%) was largely attributed to the changes in the proportion of married women during the period and reduction in the duration of postpartum insusceptibility. In the second-half of the decade, 2005 to 2011, a decline in fertility in all three categories of women’s education was observed. The decline in fertility observed among women with primary level of education was the largest (−29.7%) and all the proximate determinants played a role albeit, in varying degrees. The overall decline in fertility witnessed during the 2000–2011 period by level of education of women was highest for women with secondary and above level of education (−20.9%) and least for women with primary level of education (−12.0%).The predicted total fertility rate decline of 25.5% between 2000 and 2011 was decomposed into a 29.3% decline due to an increase in contraceptive practice and a 1.9% decline due to a decrease in the proportion of women in union, and a 9.0% increase due to shortening of the duration of postpartum infecundibility. The change in postpartum insusceptibility contributed the increase in the total fertility rate cancelling out a substantial part of the potential impact of contraception and contributing to a stall in fertility decline. Consistent with this finding, Nasir and Sinha reported the fertility-inhibiting effect of natural fertility control being gradually replaced by contraceptive use .The index of postpartum insusceptibility declined in the values of its indices in the survey periods signifying a decrease in traditional cultural practices of postpartum abstinence. Contrarily, Ngalinda and Teklu et al. found an increase in the effect of postpartum infecundability in controlling fertility overtime and a primary factor for reducing the prevailing levels of fertility compared to its biological maximum , 28.The model also confirmed that postpartum insusceptibility due to breastfeeding had the greatest inhibiting effect on fertility among illiterate women and women residing in rural areas. In societies where breastfeeding is generally prolonged and universal and contraceptive use is rare, the primary determinant of birth interval is the duration of breastfeeding –31. HoweThe fertility-constraining power of contraception showed remarkable increase overtime. Fertility decline in Africa initially began in Kenya, Botswana, and Zimbabwe due to high levels of contraceptive practice, ranging between 27 and 44% among currently married women of reproductive age . SlowingContraception led to a remarkable change in inhibiting fertility among illiterate and rural women between 2000 and 2011. This may be due to the deployment of health extension workers since 2003 in each Kebele . The accompanying diffusion effects brought by such new endeavours are contributing factors in shaping the contraceptive use and fertility behaviour of the society, especially in rural areas of the region .The index of marriage contributed a minor proportion of the decline in the total fertility rate in the region due to the small increase in age at first marriage, as marriage is a customary and socially established practice that has been carried on for generations in most communities of the region. However, the proportion of marriage is one of the major fertility-inhibiting factors and played increasing role overtime , 45–49.The predicted total fertility rate of the urban areas in the region consistently increased during the period due to a reduction in age at first marriage. In Amhara National Regional State, there has been a decline of more than two years in the median age at first marriage from 22.9 to 20.6 years in urban areas especially amongst the youngest women –16. ConsThis study has a number of strengths. First, the study utilized a large population-based sample, a randomly selected population, a high response rate (>98%), the use of standardized surveys, the well-tested validated questionnaires, and the high quality of data due to extensive training of data collectors and support during the fieldwork, with concurrent data entry and editing with feedback during fieldwork. Second, calculations were done after the data were weighted for the sampling probabilities and non-responses. Third, decomposition of proximate determinants was applied to understand the sources of change in fertility overtime. Finally, the study used a five-year interval surveys and can provide important data for monitoring population/reproductive health policies and family planning programs.Our study also has some limitations. During decomposition analysis, the index of induced abortion was taken as 1 in all populations, that is, induced abortion was assumed to have no inhibitory effect on fertility of a woman because of a lack of data in each survey. The exclusion of abortion in the model could have an effect on the estimates of total fecundity especially in areas where it is high. It is documented that induced abortion rates in sub-Saharan Africa are growing very fast, particularly in urban areas and amongst the youngest –57. AnotAn increase in the level of contraceptive use and effectiveness overtime among fertile women was the single most important contributing proximate determinant to the recent fertility decline in the region. Postpartum insusceptibility contributed to an increase in fertility due to a shortening of the duration of postpartum infecundibility during the last decade. Regardless, this increase was compensated by the fertility-inhibition effect of contraceptive use in Amhara National Regional State. Marriage contributed a little in constraining fertility in the 11-year span due to the unchanging of age at first marriage at early age (before age 18) in the region.Although fertility declined overtime in the region, a close look at fertility experience of women disaggregated by various characteristics revealed that those who live in rural areas and had primary or no education were the ones with high fertility. Fertility transition in the region could be accelerated through significantly increasing girls’ education beyond the primary level of education and strengthening education and communication about the status of women. On the other hand, strengthening of existing reproductive health and family planning programs is required in order to increase the quality and quantity of contraceptive use and achieve higher use effectiveness which will lead to a greater contribution to fertility decline. In this regard, strengthening the Health Extension Workers, Health Development Armies and other volunteers’ community health workers could have a powerful role in reaching women with family planning messages, and impacts the ability to maintain the current impetus in contraceptive use. There is the high level of unsatisfied demand for fertility regulation and the increasing contraceptive use in the future particularly in rural areas would have a major impact. In addition, the implementation of the family law which asserts the minimum age of marriage as 18 years should be ensured by creating community awareness using social mobilizations, strengthening local enforcements, and establishing legal age assessment committees in each rural Kebeles.


Exposure to sustained high concentrations of HCFC-123 is known to be hepatotoxic. We report two simultaneous cases of toxic hepatitis related to exposure to 2,2-dichloro-1,1,1-trifluoroethane (HCFC-123), a common refrigerant, at a Korean fire extinguisher manufacturing facility.Patients A and B were men aged 21 and 22 years, respectively, with no notable medical histories. They had recently started working for a manufacturer of fire extinguishers. During the third week of their employment, they visited the emergency center of a general hospital due to fever, lack of appetite, and general weakness. At the time of their visit, they were suspected as having hepatitis due to elevated aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), gamma-glutamyl transferase (GGT), and total bilirubin levels and were hospitalized. However, as their condition did not improve, they were moved to a tertiary general hospital. After conservative treatment, one patient improved but the other died from acute hepatic failure. Assessments of the work environment showed that the short-term exposure levels of HCFC-123 for valve assembly processes were as high as 193.4 ppm. A transjugular liver biopsy was performed in patient A; the results indicated drug/toxin-induced liver injury (DILI). Given the lack of a medical history and the occupational exposure to high levels of HCFC-123, a hepatotoxic agent, the toxic hepatitis of the workers was likely related to HCFC-123 exposure.Work environment assessments have not included this agent. To the best of our knowledge, we are the first to report a case of death related to HCFC-123-induced liver damage. Our findings suggest that exposure standards and limits for HCFC-123 must be developed in Korea; work environments will have to be improved based on such standards. It may cause irritation to the eyes . HCFC-12Several cases of liver damage related to HCFC-123 exposure have been reported  . The BelWhile there are no antidotes for the liver damage related to HCFC-123 exposure, conservative treatment generally leads to recovery .Fever, chills, and myalgia.Hypertension/Diabetes mellitus/Tuberculosis/Hepatitis (−/−/−/−).Social history: non-smoker, non-drinker.Medications: Finasteride, from January 2017.Patient A showed symptoms of fever, chills, and myalgia starting on August 12. The symptoms continued, and the patient visited the emergency room of a general hospital one day later. While he underwent conservative treatment, he showed over 2000 IU/L of AST and ALT and no signs of improvement; therefore, he was moved to a tertiary general hospital, together with patient B. Laboratory examinations at the tertiary general hospital on August 15 indicated clearly elevated liver enzyme levels . Conservative treatment was provided. Anti-HAV IgM, HBsAg, HBcAb IgM, anti-HBs, and anti-HCV were negative, while anti-HAV IgG was positive, likely due to a past type A hepatitis vaccination. He tested negative for human immunodeficiency virus. Because two individuals working at the same facility showed symptoms concurrently, an occupational disease was suspected. To obtain an accurate occupational history, patient interviews and material safety data sheet checks were conducted. It was suspected that HCFC-123 was the most likely cause of hepatotoxicity. The patient underwent a transjugular liver biopsy that indicated portal and panlobular infiltration of mixed inflammatory cells (abundant eosinophils) with lobular disarray, multifocal spotty and confluent necrosis, mild bile duct damage, and minimal macrovesicular and microvesicular steatosis; these findings were considered suspicious for DILI as they demonstrated patterns of acute hepatitis. The patient’s liver enzyme levels rose to AST 3108 IU/L and ALT 3265 IU/L but then decreased on August 21 to AST 117 IU/L and ALT 662 IU/L. Subsequently, he was improved and moved to a general hospital. The progression of patients A and B were shown in Tables Fever, anorexia, and general weakness.Hypertension/Diabetes mellitus/Tuberculosis/Hepatitis (−/−/−/−).Social history: smoker (4 pack-years), social drinker (4 standard drinks per week).Patient B reported fever, anorexia, and general weakness starting on August 11. As the symptoms continued, he visited the emergency room of a general hospital 2 days after the onset of the symptoms. Upon admission, laboratory examinations indicated thrombocytopenia and the following findings: hemoglobin 17.4 g/dL, leukocytes 7500/μL, platelets 94,000/μL; blood urea nitrogen (BUN) 17.5 mg/dL, and creatinine 1.14 mg/dL. Elevated liver enzyme levels were found . The prothrombin time showed an international normalized ratio (INR) of 1.35, indicating slight prolongation. The serology tests that were conducted due to the high liver enzyme results showed that the patient tested negative for anti-HAV IgM, anti-HAV IgG, HBsAg, HBcAb IgM, anti-HBs, and anti-HCV. Routine urinalysis showed signs of proteinuria. The patient was hospitalized and underwent conservative treatment. However, as he did not show any improvements but presented with elevated liver enzyme levels, thrombocytopenia, and acute renal failure, he was moved to a tertiary general hospital 2 days later. The laboratory examinations at the tertiary general hospital showed the following: AST 13,537 IU/L, ALT 7115 IU/L, GGT 155 IU/L, and total bilirubin 3.6 mg/dL. The patient underwent tests to identify the cause of the hepatitis and to rule out infections. IgM anti-HEV and IgG anti-HEV were both negative; thus, infection by the hepatitis E virus was considered unlikely. He also tested negative for human immunodeficiency virus, cytomegalovirus, Epstein-Barr virus, herpes simplex virus, and varicella-zoster virus; thus, a recent infection was excluded. IgG for measles and mumps was positive, and IgG for rubella was in the borderline positive, which was likely caused by a recent measles, mumps, and rubella (MMR) vaccination. Antibody tests for the Hantaan virus, Leptospira, Rickettsia typhi, and Rickettsia tsutsugamushi were negative. A stool culture did not isolate salmonella, shigella, or vibrio species, and no growth was detected in a blood culture. Tests for autoimmune hepatitis showed positive antinuclear antibody staining with a speckled pattern at 1:40. Anti-mitochondrial, anti-neutrophil cytoplasmic, smooth muscle, and anti-LKM antibodies were all negative. The patient’s serum IgG concentration was 530 mg/dL, which is below the reference range (700–1700 mg/dL). Therefore, the patient likely had no autoimmune hepatitis. No liver biopsy could be performed as the patient had a poor general condition due to a cerebral hemorrhage. However, a transjugular liver biopsy could be performed in his co-worker (patient A) who was co-hospitalized; these results indicated drug/toxin-induced liver injury (DILI). Therefore, it appears likely that patient B also had DILI. On August 17, he showed a lowered level of consciousness, hypersensitivity, and prothrombin time prolongation; therefore, he was moved to the intensive care unit (ICU) under suspicion of acute fulminant hepatitis and underwent continuous renal replacement therapy and Molecular Adsorbent Recirculating System (MARS®) therapy. There is no specific treatment for HCFC-123-induced toxic hepatitis. Although an occupational disease was suspected, and the patient was referred to an occupational and environmental medicine physician, the disease progression could not be slowed. The patient did not respond to conservative treatment. Seven days later, on August 24, the patient died from acute hepatic failure while undergoing treatment at the ICU.Patient A began work at the fire extinguisher manufacturing facility on July 26; his symptoms of fever, chills, and myalgia began 17 days later on August 12. Patient B started working at the same facility on July 27; his symptoms of fever, anorexia, and general weakness began 15 days later on August 11. The company manufactures fire extinguishers and gas sprayers and uses HCFC-123, HFC-125, HFC-236FA, and loaded stream. HCFC-123 charging involves a simple procedure, and both, contract workers and new employees are in charge of this process. The fire extinguisher manufacturing process consists of component assembly prior to charging, charging of the HCFC-123 fire extinguishing agent, packing, and consignment. Typically, the patients A and B were tasked with component assembly, although they were often involved in the entire manufacturing process. The HCFC-123 charging process involves filling the storage tank with HCFC-123, placing the empty fire extinguisher on the conveyor belt, charging the empty extinguisher cans with HCFC-123, assembling the valves and seal rings on the fire extinguishers, nitrogen charging, and manual sealing tests. Filling the empty fire extinguisher cans with HCFC-123 is automated as they move across the conveyor belt; the charging time per extinguisher is approximately 20 s. During the first two weeks of employment, the patients A and B were not tasked with HCFC-123 charging duties. Patients A and B were tasked with HCFC-123 charging for about 5 h split up into 3 days between August 9 and August 11. Patient A was tasked with nitrogen charging, whereas patient B was involved in the valve assembly process.The work environment was assessed for the time-weighted average (TWA), short-term, and area exposure levels of HCFC-123 Tables . The TWAIn both, short-term and area exposure assessments, HCFC-123 levels were highest for the valve assembly process, likely because this process was in close physical proximity (about 1 m) to the automatic HCFC-123 injector. In contrast, nitrogen charging was performed about 3 m away from the automatic HCFC-123 injector.We report two cases of liver damage due to exposure to HCFC-123 in workers working at a fire extinguisher manufacturing facility in Korea. Patient B died, whereas patient A recovered. Whereas patient B performed valve assembly during the HCFC-123 charging process, patient A was tasked with nitrogen charging. HCFC-123 exposure levels were highest for the valve assembly process, which is in closer proximity to the automatic HCFC-123 injector, in both, short-term and area exposure assessments. Therefore, patient B was likely exposed to higher levels of HCFC-123 than patient A, and this higher exposure likely led to his death. The workers were not familiar with the risks related to HCFC-123, which may have affected their behavior. An automatic charge line was implemented at the facility in April of the same year, followed by an increase in production, making indirect exposure to HCFC-123 possible. Therefore, health examinations were conducted for all co-workers in the same workplace. In these health examinations, 6 of 15 co-workers showed abnormal liver tests. Moreover, a further investigation revealed one patient who was suspected of having toxic hepatitis in June of the same year.No exposure limits for HCFC-123 have been set in Korea. In the US, the AIHA Workplace Environmental Exposure Levels (WEEL) Committee set the TWA exposure limit at 50 ppm, based on a 40-h workweek , 14. As To date, it has been thought that hepatotoxicity can be avoided if the 8-h TWA exposure limit of 50 ppm set by the AIHA WEEL Committee is met and safety equipment is worn by workers. However, in this case the 8-h TWA for the workers was 19.1–20.9 ppm. As Korea has set no exposure standards for HCFC-123, defining such exposure standards and limits should be prioritized.The diagnostic criteria for DILI are an ALT level of > 5 times the upper limit of normal (ULN), or an ALP level of > 2 times the ULN, or an ALT level of > 3 times the ULN with a bilirubin concentration of > 2 times the ULN . There a4 and argon. HCFC-BLEND B consists of > 93% HCFC-123 [HCFC-123 is used globally, especially in the form of HCFC-BLEND B by mixing CFHCFC-123 . When usHCFC-123 , 22. ThePrevious cases of HCFC-123-induced liver damage were related to HCFC-123 being used as a refrigerant or solvent; moreover, there have been no reported deaths so far. In this report, HCFC-123 was used as a fire extinguishing agent by workers at a fire extinguisher manufacturing facility. It has been reported that HCFC-123-induced liver damage can be effectively treated with conservative treatment; however, a death occurred in our case. The concentrations of HCFC-123 were not higher in this case when compared to past cases. There is a possibility that host idiosyncrasy might affect the liver in patient B.As HCFC-123 is widely used as a fire extinguishing agent, its safety should be reevaluated. To reduce the occupational risks of HCFC-123 exposure, it is crucial to maintain a health & safety management system, foster technological innovation, adhere to regulations, and implement effective employee oversight strategies .In conclusion, there have been only few cases of liver damage from HCFC-123 exposure globally. To the best of our knowledge, we are the first to report a case of death caused by HCFC-123-induced liver damage. Conservative treatment has been thought to be effective in the treatment of CFC-123-induced liver damage; however, in our case, one patient died. Unlike previous cases, exposure to HCFC-123 happened at a manufacturing facility for fire extinguishers in this case. Our findings indicate that exposure standards and limits should be set in Korea.