Background: Disease prioritization aims to enhance resource use efficiency concerning human and animal health systems’ preparedness and response to the most important problems for the optimization of beneficial outcomes. In sub-Sahara Africa (SSA), several prioritizations of zoonoses and transboundary animal diseases (TADs) have been implemented at different scales to characterize potential disease impacts. Method and principal findings: In this systematic review, we analyze the methodologies used, outcomes, and their relevance by discussing criteria required to align decision-makers’ perceptions of impacts to those of other stakeholders for different prioritization in SSA. In general, the sectorial representativeness of stakeholders for processes implemented with the support of international partners showed slight differences with the absence of local stakeholders. Whatever the tool prioritized, zoonoses were similar in general because of the structured nature of those tools in assessing decision-makers’ preferences through value trade-offs between criteria while ensuring transparency and reproducibility. However, by involving field practitioners and farmers, there were different outcomes with processes concerning only decision makers and experts who were more sensitive to infectious TADs, while the former raised parasitic disease constraints. In this context, multicriteria decision analysis-based zoonoses and TADs prioritizations involving a balanced participation of stakeholders might contribute to bridging these divergences, whatever the scale. Conclusion and significance: Prioritization processes were important steps toward building and harmonizing technical laboratory and surveillance networks to coordinate projects to address priority zoonoses and TADs at the country and/or sub-regional level. Those processes should be enhanced. The use of prioritization exercises for the ranking of diseases has spread from America (Canada) in 1987 . PrioritInternational bodies such as World Health Organization (WHO), World Organization for Animal health (OIE), and Food and Agricultural Organization (FAO) attach importance to the prioritization of diseases. They implemented international disease prevention and control strategies and proposed disease prioritization as a way to improve various diseases prevention and control efforts. In this regard, in 2006, the WHO published guidelines on communicable disease surveillance priorities that emphasize a prioritization role in improving disease surveillance systems at national and regional levels ; then, tThe recent growing interest and advocacy by non-profit organizations, governments, industries, and academia toward the “One Health Approach” in mitigating overlapping health problems between human, animal, and environment sectors have triggered the prioritization of zoonotic diseases in several low–middle income countries worldwide in order to coordinate preparedness and response across sectors more effectively . A recenFour countries  implemented both animal and zoonotic disease prioritization processes at the national level using the Phylum and the OHZDP tool;Six countries  were concerned only by zoonotic disease prioritization at the national level using the OHZDP tool;Six countries  used the Phylum tool to prioritize TADs and zoonoses at the national level.The database search yielded nine relevant animal prioritization reports and 16 published papers relevant to our study. All studies were carried out and published between 2015 and 2021 . Three aA local prioritization of animal diseases (participatory epidemiology) concerning one of the sixteen countries above (Ethiopia) and regional prioritizations of zoonoses and animal diseases in West and East Africa were retrieved as well.For regional and national prioritization, workshops were specifically conducted, and all processes aimed at identifying and prioritizing either only zoonotic diseases or both zoonoses and animal diseases. Subject expert matters (or trained facilitators) contributed to the processes whatever the tool used following standard procedures previously described. During the OHZDP process at the national level, participating members were grouped by their agencies and voted on the ranking or weight applied to each criterion before conducting a final ranking of diseases. Participants belonged to different national sectorial or administrations including public health , animal health , wildlife and environmental health , research institutions , local universities , international partners, one health coordinating institutions (one-third of the countries), and military and security services (one-fifth of the countries). The private sector was involved only in two out of the ten concerned countries, and the participants were from central or national administration for all countries . The parThe prioritization of TADs and zoonoses using the Phylum tool was carried out by senior experts from the national veterinary departments , public health departments (three-fifths of the countries), and international partners . Wildlife, administrative authorities, and One Health coordinating institutions were each involved in only one of the ten countries. In all countries, participants were mainly from the public sector and from central or national administration for all countries .Severity of disease in humans ,Availability of interventions  ,Economic, environmental, and/or social impact (9/10 countries),Presence of disease in country and/or region (8/10 countries),Epidemic/pandemic potential (and/or sustained transmission in humans) (7/10 countries).For the OHZDP process, irrespective to the assigned weight, the most frequent disease ranking criteria used were :SeverityWith the Phylum tool, the importance of the listed diseases was assessed based on the scoring of selected specific criteria including local economic impact, local public health impact, local societal impact, local environmental impact, and the local feasibility, economic, societal, and environmental impact of control measures. The assessments considered both present and absent diseases (Phylum). For each listed disease, the tool uses expert opinion bases on data from public health and veterinary services, statistics from health information systems  from OIE, FAO (Empres-I), AU/IBAR (Aris) OMS) for the prioritization of TADs and zoonosis. The analysis of a disease is made in two sequential steps: first, a global characterization of the disease, aiming at assessing the different above-mentioned impacts within the region, independent of any particular local context . Dependin = 10), anthrax (n = 7), brucellosis (n = 7), zoonotic influenza virus (n = 7), and hemorrhagic fever (Ebola/Marburg) (n = 6) were the top five most prioritized zoonoses amongst the studied countries. Of that ranking zoonotic influenza virus (n = 2), hemorrhagic fever (Ebola/Marburg) (n = 2), trypanosomiasis (n = 1), and dengue (n = 1) were the ones ranked as exotic zoonotic diseases. Within the ECOWAS region, 30 zoonotic diseases were listed, and seven , zoonotic influenzas, zoonotic tuberculosis, trypanosomiasis, and yellow fever) were selected as priority zoonoses. Eleven, five, and two zoonoses prioritized by the ten countries were caused by viruses, bacteria, and parasites, respectively. As a result of the OHZDP tool’s ranking process, the ten countries listed on average 38.5 zoonotic diseases (range: 11–48) of which 5.7 (range: 5–7) and 0.6 (range 0–1) endemic and exotic zoonoses were prioritized, respectively . Eighteen = 10), peste des petits ruminants (n = 9), contagious bovine pleuropneumonia (n = 8), New Castle disease (n = 7), and contagious caprine pleuropneumonia (n = 5). Brucellosis (n = 8), rabies (n = 7), and tuberculosis (Mycobacterium bovis) (n = 6) were zoonoses ranked by the most countries. In addition, exotic priority diseases were ten TADs and four zoonoses where specific serotypes of foot and mouth disease (n = 9) was the main exotic TADs ranked (n = 10) and Rift valley fever (n = 5) were the most prioritized zoonoses when being exotic to the country. Endemic TADs prioritized were caused by viruses (n = 6), bacteria (n = 2), and parasites (n = 1). Three, four, and two endemic zoonosis were caused by viruses, bacteria, and parasites, respectively. Eight and two exotic TADs prioritized were caused by viruses and bacteria, respectively. All exotic zoonoses (n = 4) prioritized were caused by viruses.Amongst the ten countries that implemented the Phylum prioritization process, 15.90 (range: 11–23) of which 8.80 (range: 6–12) and 3.50 (range 2–6) were endemic and exotic TADs and zoonoses were prioritized respectively in each country . Nine ens ranked . Highly n = 16) using previously developed prioritization tools. In addition, regional (West and East Africa countries) and local (District) level prioritization processes were also taken into consideration. To the best of our knowledge, this study is one of the few of its kind to gather knowledge generated through several prioritization processes using different tools in SSA whereas previous synthetic works focused on analyzing data generated through the processes using the same tool [The need of achieving the optimal benefit in tackling animal and human health challenges or constraints has prompted the prioritization of diseases as a strategic process ensuring efficient use of limited resources to target the most important problems. The process of prioritizing animal diseases and zoonoses has been implemented previously in different locations, with different magnitudes and purposes in South and North America ,38, Euroame tool . For the prioritization of animal diseases and zoonoses, it has previously been argued that using an interdisciplinary team of participants (facilitating and voting) who stay neutral, unbiased, and do not focus on their specific sector, affiliation, or area of expertise enables voting members’ voices to be heard and recognized ,11. In tWhatever the tool used at the country level (CDC OHZDP or OIE Phylum), zoonotic diseases that were most frequently prioritized were rabies, brucellosis, hemorrhagic fevers (Marburg/Ebola), anthrax, Rift Valley fever, and zoonotic tuberculosis. For public health, this consensus on prioritized zoonoses also shows how these endemic diseases are important at the country and continental level. Rabies is an acute and progressive viral encephalitis causing approximately 59,000 human deaths annually in Africa and Asia ,46, and In total, four parasitic diseases  were ranked between the 18 top diseases, suggesting that decision makers and experts’ perception of the impact of bacterial and viral infections is higher than the one of parasitic diseases. However, developed countries that have succeeded in controlling most of the diseases prioritized in Africa are mainly focused on the introduction of emerging and re-emerging infectious and food-borne  diseases ,41,42,73At the time of this study, five countries  that prioritized zoonotic diseases using the OHZDP tool had not published their reports yet, and all Phylum country prioritization processes were considered. Thus, the prioritization processes reviewed in this study and carried out at the regional and national level are in no way exhaustive though representative and may be only partly useful at the local level where a full list of disease constraints should be identified based on production systems. In this regard, prioritization processes should be implemented in a standardized format that can be flexibly used whatever the scale and stakeholders involved such as multicriteria decision analysis (MCDA), which is a set of methods from decision science that have been recently used for disease prioritization ,77. HoweWe attempted to gather details of all published  studies and reports involving animal and zoonotic disease prioritization in sub-Sahara Africa (SSA) . CountryAnimal husbandry in SSA is strongly constrained by diseases that have various economic and public health impacts. In this critical review, we have analyzed several zoonosis and animal disease prioritization processes carried out in SSA. By aggregating data for tools that were used several times for prioritization, we characterized criteria and participating stakeholders. Then, diseases that have the greatest impact on livestock production and public health according to stakeholders were identified and ranked. There was a general agreement of priority zoonoses whatever the tool used for prioritization except for some diseases or syndromes seen as important for farmers or practitioners. The limitations of each prioritization process were discussed and ways of improvement were presented. Few research studies have focused on synthetizing disease prioritization outcomes from diverse prioritization methods and questioning the operationalization of the surveillance and control of the prioritized diseases. Thus, in SSA, the operationalization of outcomes from the prioritization of zoonoses and TADs at the regional, country, local, and farm levels can be certainly improved by involving all stakeholders particularly at the local level through standardized prioritization methods such as multicriteria decision analysis.


The second most common cancer among females in Bangladesh is cervical cancer. The national strategy for cervical cancer needs monitoring to ensure that patients have access to care. In order to provide accurate information to policymakers in Bangladesh and other low and middle income countries, it is vital to assess current service availability and readiness to manage cervical cancer at health facilities in Bangladesh.An interviewer-administered questionnaire adapted from the World Health Organization Service Availability and Readiness Assessment Standard Tool was used to collect cross-sectional data from health administrators of 323 health facilities in Bangladesh. Services provided were categorized into domains and service readiness was determined by mean readiness index (RI) scores. Data analysis was conducted using STATA version 13.There were seven tertiary and specialized hospitals, 118 secondary level health facilities, 124 primary level health facilities, and 74 NGO/private hospitals included in the study. Twenty-six per cent of the health facilities provided services to cancer patients. Among the 34 tracer items used to assess cancer management capacity of health facilities, four cervical cancer-specific tracer items were used to determine service readiness for cervical cancer. On average, tertiary and specialized hospitals surpassed the readiness index cutoff of 70% with adequate staff and training (100%), equipment (100%), and diagnostic facilities (85.7%), indicating that they were ready to manage cervical cancer. The mean RI scores for the rest of the health facilities were below the cutoff value, meaning that they were not prepared to provide adequate cervical cancer services.The health facilities in Bangladesh  lack readiness in cervical cancer management in terms of guidelines on diagnosis and treatment, training of staff, and shortage of equipment. Given that cervical cancer accounts for more than one-fourth of all female cancers in Bangladesh, management of cervical cancer needs to be available at all levels of health facilities, with primary level facilities focusing on early diagnosis. It is recommended that appropriate standard operating procedures on cervical cancer be developed for each level of health facilities to contribute towards attaining sustainable developmental goals. Over the years, the increasing burden of non-communicable diseases (NCDs), such as diabetes, hypertension, cardiovascular diseases, and cancer has become a significant global public health concern , 2. StudThe second most predominant type of cancer among women in LMICs is cervical cancer, next to breast cancer , 8. EighLike other LMICs, cervical cancer is the second most common cancer among females in Bangladesh , with 8068 new cases and 5214 deaths caused by it every year –13. The With scientific advancement and changing health dynamics, health systems researchers have identified critical areas that can improve cancer management in developing countries if addressed . These iAt this juncture of mid-way in the implementation of the National Strategy for Cervical Cancer Prevention and Control Bangladesh (2017–2022), an assessment of the program implementation using the Service Availability and Readiness Assessment (SARA) Survey may be of value to note the achievements, and highlight the areas where further strengthening or updates in line with current scientific research may be needed . The stuIt was a cross-sectional sub-study of a more extensive parent study called Service Availability and Readiness Assessment (SARA) Survey for NCDs and Disability Service Delivery System in Bangladesh. The study was conducted from December 2017 to June 2018. In this sub-study, variables for assessing the availability and readiness of cancer-related services  were considered.WHO SARA Standard Tool was used to collect data on the availability of medical equipment related to cancer including their location, functional status, and components of support systems  .A list of all available public and private health facilities was compiled at the primary, secondary and tertiary levels in Bangladesh using the Management Information System (MIS) and and hospital administration records from the Directorate General of Health Services (DGHS); the National Institute of Population Research and Training (NIPORT) and from the Bangladesh Health Facility Survey (BHFS) 2014 , 18. TheOf these 19,184 health facilities, the parent study of SARA Survey for NCDs and Disability Service Delivery System in Bangladesh selected a sample of 590 health facilities at various levels. These included ten randomly selected tertiary and specialized hospitals for NCDs . All secondary level health facilities, that is, 60 district hospitals (DHs) and 61 Mother and Child Welfare Centre (MCWCs), were also included in the sample. Among the primary level health facilities, the sample also included 128  Upazila Health Complexes (UHCs), and 256 Family Welfare Centres (FWCs) and Community Clinics (CCs). Further, we included randomly selected 75 NGOs/ private hospitals.This sub-study then excluded 267 health facilities from analysis, which included specialized hospitals that did not focus on cervical cancer care  and facilities where a staff member was not available to complete the survey. The study included a final sample size of 323 health facilities, as depicted in Fig. The head of each hospital or a management staff member who would have sufficient knowledge of hospital capacity and operations were selected to participate in the survey Table .Table 1Forty-eight interviewers, all Bachelor of Medicine and Bachelor of Surgery (MBBS) graduates, were trained for 2 days on the data collection instrument and data entry into RedCap software using tablets. The interviewers received instructions for uploading the data immediately to the server from each health facility, enabling researchers at the head office to monitor the data collection in real-time. The health facilities included in this study were contacted beforehand and an appointment was set for the interviews. The interviews were conducted in-person and recorded.Descriptive analyses were performed based on the dataset using the standard core indicators  and corresponding tracer indicators, as per the reference manual of SARA . There wTwo outcomes, the “availability” and “readiness” of cancer services, were calculated based upon previous studies that utilized the SARA tool , 21. SerData was collected on the availability of any type of cancer service readiness tracer indicators at all health facilities.Tracer indicator scores were calculated for all cancer tracer indicators at each health facility level, using the following formula: number of health facilities with individual tracer indicators present *100/total number of health facilities.Next, the tracer indicator scores specifically for cervical cancer were extracted  which fell under three domains . Two of the indicators, guidelines and training, covered the domain of staff and training. Each of the other two indicators covered the domains of equipment and diagnostic tool respectively.In the next step, the cervical cancer-specific readiness index (CSRI) scores for the cervical cancer-specific domains were calculated for each health facility level. CSRI was done by determining the mean of all tracer indicator scores for each domain (sum of tracer indicator score in each domain/number of tracer indicators in each domain) for each facility level.The mean cervical cancer-specific readiness index (mean CSRI) score was calculated at each level of health facility by determining the average of the CSRI score of all three domains (sum of readiness index in each domain/number of domains) at each facility level.Cervical cancer-specific readiness index (CSRI) scores and mean index scores were then compared to a cutoff level of 70%, as evident from a study in Zambia that also used the SARA tool and considered health facilities to be “ready” to manage NCDs when their mean readiness index scores were above the same cutoff score . All anaA total of 323 health facilities were assessed, where 248 (76.8%) were public and 75 (23.2%) were from the private sector. Facilities included 124 UHCs (primary level) (38.4%), 74 private/NGO hospitals (22.9%), 58 DH (secondary level) (18.0%), 60 MCWC (secondary level) (18.6%), and 7 tertiary level specialized hospitals (2.2%). Most of the NGO and private hospitals are secondary level health facilities. However, only one of the tertiary level specialized hospital was a private hospital. More than one-fourth of facilities were situated in the Dhaka division (Table Almost three-quarters of the facilities (74%) did not provide any service to cancer patients Fig. . All sevFrequencies and tracer indicator scores for cancer at each health facility level.The tracer indicator scores for all 34 cancer tracer indicators at each level of health facility are provided in Table The tracer indicator scores specific for cervical cancer were 100.0% for guideline, training, and speculum and 85.7% for acetic acid at the tertiary and specialised hospitals. The tracer indicator scores specific for cervical cancer was < 18% for guideline and training, and 62.1% for speculum and acetic acid at the secondary level of health facility, while that at the primary level of health facilities as well as the NGO and private hospitals was < 15 and < 50% respectively.Cervical cancer-specific readiness index (CSRI) scores at each health facility level: Fig. Mean cervical cancer-specific readiness index (mean CSRI) scores at each health facility level: Fig. This study identified several gaps that require intensive attention by policymakers and implementer to attain the desired targets. During the study, information regarding availability of staff and guidelines, screening equipment, diagnostic facilities and medicines were collected. While issues were relatively good at the tertiary level, more needs to be done at other levels of health facilities. This study also demonstrated that only tertiary-level facilities in Bangladesh are ready to diagnose and treat for cervical cancer patients while only some facilities at the primary and secondary levels have adequate availability and readiness for diagnostic services.In Ethiopia, a study found that the availability and management of cervical cancer among NCD service providing health facilities was almost negligible, while little over half of the health facilities had trained staff for cervical cancer , 22. StuUsing the SARA tool to assess service availability and readiness for major non-communicable diseases, a study conducted among health facilities in a selected district of Bangladesh found that service availability was less than 40%. The overall readiness score, though variable for different diseases, was close to 70% or below it, indicating that the health services strengthening are warranted to meet the targets set [The finding gives the impression that the implementation of the Bangladesh Cancer Control Strategy, which came into effect in 2017 has a long way to go to reach the desired goal by 2022. Less than half of the primary level health facilities and just over half of the secondary level facilities were prepared to diagnose cervical cancer. But in the absence of trained human resources and guidelines, these facilities cannot be brought into action for rendering the desired services. As early detection of cervical cancer is the key to favourable treatment outcome, diagnostic facilities through training and guidelines or standard operating procedure (SOP) need to be strengthened at the primary level. The existing referral mechanism needs to be further streamlined so that the cervical cancer patients can avail treatment at secondary or tertiary level health facilities without delay . The secThis study’s strengths included health care facilities from rural to urban under public and/ or private support and explored a large representative sample size. On the other hand, since this was a cross-sectional study and the data collected was only valid for the time when the hospital heads were interviewed. Therefore, no trend or change over time could be witnessed in this study. Also, the study could not include all the sample health facilities due to incomplete questionnaire and absence of proper interviewee. While data on the availability of staff and guidelines, equipment for screening, diagnostic facilities and medicines were collected, information on area specific population density was not collected, which was why bed density, facility density, health worker density, and so forth could not be estimated. Further studies to explore the state of HPV vaccine for the targeted population as per national strategy is needed. While working with the SARA tool, the method has some limitations, specially, for cervical cancer services. The SARA tool allowed data collection on the availability of tracer items relevant to VIA, but not on those relevant for other diagnostic methods such as thermocoagulation and cervical conization. Diagnostic and services for cancer are multidisciplinary requiring specific specialization, an issue not adequately focused on the SARA tool to assess the availability and readiness , 25. As The study findings suggest that health facilities in Bangladesh  lack readiness in cervical cancer management in terms of guidelines on diagnosis and treatment, staff training, and shortage of equipment. Given the high prevalence of cervical cancer in Bangladesh, such cervical cancer management needs to be available at all health facilities levels. If probed, evidence of similar situation may be found in other LMICs too. This study was a basic assessment of service availability and readiness. Further studies can be conducted to collect data on machines for cryotherapy, thermocoagulation and LEEP, capacity to perform cervical conization, hysterectomy and radical hysterectomy, capacity for cancer-related imaging, and capacity for external-beam radiation therapy, brachytherapy, and platinum-based chemoradiation. Alongside this, a well-designed plan to ensure the availability of adequate and competent care services for cervical cancer at public and private sectors following other SARA modules, such as quality of care and safety and management and finance, is needed. It may be appropriate for policymakers to develop standard operating procedure on cervical cancer to guide the care services to achieve the set targets and contribute towards attaining sustainable developmental goals.


The UK can be proud of the fact that numerous native breeds of sheep have been developed here that possess unique phenotypic features and excellent productivity and are utilized throughout the world. Their remarkable popularity and further sustainable breeding on grass pastures of British Isles and elsewhere can benefit from genomic applications.Sheep farming has been an important sector of the UK’s economy and rural life for many centuries. It is the favored source of wool, meat and milk products. In the era of exponential progress in genomic technologies, we can now address the questions of what is special about UK sheep breed genotypes and how they differ genetically form one another and from other countries. We can reflect how their natural history has been determined at the level of their genetic code and what traces have been left in their genomes because of selection for phenotypic traits. These include adaptability to certain environmental conditions and management, as well as resistance to disease. Application of these advancements in genetics and genomics to study sheep breeds of British domestic selection has begun and will continue in order to facilitate conservation solutions and production improvement. Sheep are an integral element of UK countryside landscapes at a range of altitudes, with both modern and ancient breeds  represented . Their uThe UK is a largest producer of sheep than any EU country, counting nearly 34 million adult animals and lambs . Prior tA glance at the gene pool landscape for British sheep breeds from the National Sheep Association leads to a list of over 60 native breeds , whereasSheep breed gene pool of British Isles can be subdivided into geographical groups  accordinThe geographical concentration (endemism) of the British sheep breeds was suggested to be a major risk factor for breed endangerment .British sheep breeds are common in other parts of the world and have been exported from the UK to other countries to create new breeds and improve the extant ones, with Australian, USA and European breeds being prominent examples ,18. As iMany of British breeds are unique in terms of phenotypic traits and adaptation to local conditions  see exa but noneLaminaria [In terms of peculiar adaptability, specific phenotypes and features of the British sheep breeds, some of them are lowland breeds , some are hill breeds , and some are upland breeds , while some others are adapted to variable conditions  . An extraminaria .Body size varies from small (Black Welsh Mountain) to medium  to large . Some breeds are horned , others are hornless , and in some others only males have horns . The Border Leicester breed has a characteristic “Roman nose”.Most breeds are used for producing lamb meat, while some (Clun Forest) are dual purpose and can be used for meat, wool and milk. There are long-woolled breeds , and others  naturally shed wool, not requiring shearing. Females of some breeds  have a slightly shorter gestation period allowing 1.5 lambs a year. Some breeds  have a low disease risk. Previously, British sheep breeds and flocks were assessed, on the base of prion protein (PrP) genotypes, for risk of scrapie, also known as prion disease and transmissible spongiform encephalopathy ,33,34,35Over several decades, there has been a general trend in the world’s animal production to substitute or cross local breeds with productive commercial stocks that may lead to genetic diversity reduction . Such anThe Rare Breeds Survival Trust names 57 British native sheep breeds, including 25 breeds on the critical list, e.g., Border Leicester, Dorset Horn, Balwen Welsh Mountain, Hill Radnor and Llanwenog . Indeed,The collection of breed data including adaptation to specific habitats facilitates development of conservation solutions. Characterization of genetic originality and authenticity of a breed via its DNA analysis is a crucial component in informed decision making to decide whether a breed (or population) is worthy or not of conservation action. Information regarding the risk status and the level of inbreeding can be further used to choose respective conservation strategies . Novel molecular and advanced genomic tools can be effectively employed to incorporate genetic variability and genomic information into conservation planning and sustainable conservation programs ,41. CharCLPG) locus causing the muscle hypertrophy phenotype [At present, there is a rich arsenal of genetic and genomic resources, tools and applications used for livestock assessment, breeding and production including, first of all, genetic profiling of diverse breeds, and search for quantitative trait loci (QTLs) and candidate genes in farm animals. These genomic advances facilitate breed improvement and understanding of the genetic processes in the course of domestication and breed evolution. For example, the Dorset breed was found to be a carrier of a single polymorphism mutation at the callipyge  in three other candidate genes for nematode resistance and body weight were examined in populations of domestic Scottish Blackface and free-living Soay ewe lambs, and a nominally significant association between an IL23R SNP and body weight was found [Genetic resistance to nematode infection is an important target of selective breeding for this trait in the UK. This was studied within a purebred Scottish Blackface flock by partial resequencing genes in the Major Histocompatibility Complex (MHC) class II region . Causal as found .TMEM8B and SPAG8 that showed picks of a signature of selection at single SNPs in four sheep breeds, the Suffolk among them [Other examples of candidate genes, for example, associated with ewe mature weight are ong them .With the advent of next generation sequencing (NGS) technologies, SNP panels and a whole genome sequence draft became available for the sheep by 2010 ,60 that These state-of-the-art resources are crucial for genetic improvement of the existing sheep flock by implementing genome-wide association studies , analysAfter marker validation, genetic or genomic selection is feasible when targeting, for example, such sheep traits as footrot resistance  and matuThere are two major collaborative sheep genomics groups, the International Sheep Genomics Consortium  and an AUse of multiple SNP markers has substantially enhanced analysis of genetic diversity and population history ,72,73,74The earliest research of SNP-based diversity in UK sheep showed genetic distinctiveness of three English native hill breeds examined at three SNP loci associated with phenotypes ,50. In aSNPs have also become genetic markers of choice in searching for QTLs and conducting GWAS in sheep  and selection signatures in six commercial European meat breeds including the Suffolk sheep (of Irish population) . The SufFurther development of NGS platforms and reduction of their cost make it possible to implement whole genome sequencing for numbers of individuals within one or more species. Whole genome sequences seem to provide ultimate evaluation of genetic variability and candidate mutations that can be further used for genome-wide association studies, sequence genotype imputation and genomic prediction improvement as a component of genomic selection ,66.Using whole genome sequences of 21 Chinese native sheep breeds, Yang et al.  identifiNaval-Sanchez et al.  sequenceOn the basis of whole genome sequences for 145 wild and domestic sheep and goat samples, Alberto et al.  found seUsing SNP and whole genome sequence data for a large worldwide sample collection of wild and domestic animals, Chen et al.  found thThe ongoing Australia- and New Zeeland-based project SheepGenomesDB is aimed at sequencing 453 (Run1) and 935 (Run2) animals, which is supposed to cover a global sheep breed diversity in order to identify causative mutations and facilitate genomic selection ,66.To date, there has not been launched an overall, comprehensive genetic/genomic survey of the British sheep gene pool. Certain British native breeds have been part of either diversity studies using different molecular markers or whole genome sequences and often a limited number of samples per breed. For example, using the Illumina HiSeq 2000 platform, three individual animals, each representing one Welsh sheep breed , were sequenced in order to explore their demographic history .Recently, more whole genome sequences were generated for separate British breeds. For instance, the Illumina whole genome sequences for seven Cambridge sheep and one British du Cher individual were obtained in a comparative study including also the Romanov and two Iranian breeds . The CamWhole genome sequences of 17 Poll Dorset sheep was compared to those from two Tibetan breeds . SelectiPDGFD, XYLB, TSHR, and SGCZ, with PDGFD (platelet derived growth factor D) being a specially remarkable candidate for fat deposition in tail.There are also international projects that have generated whole genome sequences for British and non-British breeds and can be served as data sources for further studies ,89,93,94The UK sheep landscape is characterized by abundant breed history and genetic diversity, suggesting its preservation for sustainable use and advanced research. The latter can rely upon state-of-the-art molecular and genomic tools including SNPs and whole genome sequencing. This opens further opportunities to elucidate breed ancestry, variability, and genetic merit of particular markers and candidate genes for adaptation, genomic selection and production improvement. Further research is anticipated to understand comprehensively how genomic landscape of the British sheep has contributed to the fact that over the centuries they have thrived to current numbers and value, delighting our eyes on the green landscapes of today’s Great Britain.


Assessing value in child health is increasingly important as health care systems face difficult choices with regard to what services and programs for children to fund and deliver. Measuring cost-effectiveness, evaluating effects on quality of life for patients and their caregivers, determining preferences for treatments and services from patients, families, and members of the public, and consideration of the social, legal, ethical, and equity implications of new complex technologies all contribute to an understanding of value. Conducting research to measure elements of value often necessitates novel methods and approaches. Conducting such research in child health is particularly challenging given the paucity of valid pediatric patient-reported outcome instruments, the lack of rigorous effectiveness data sources, reliance on proxy reporting, and many other methodologic and data-related constraints. This Special Issue features papers exploring topics that expand our understanding of value in child health. Not surprisingly, many papers focus on the measurement of health-related quality of life (HRQoL) ,2,3,4. IUnderstanding and accurately measuring HRQoL is but one aspect of full economic evaluation. A significant challenge that has received little attention is how to convert deficits in child cognitive function into a measurable future productivity loss that can be modeled in economic evaluations of interventions aimed at improving cognitive function over the lifespan. Using published estimates of the correlation between intelligence quotient (IQ) and market productivity, Gosse and Zhou valued a one-point improvement in IQ in the United States at USD 10,600–13,100 [Lifetime time horizons are essential for economic evaluations of precision medicine screening and diagnostic testing interventions that identify genetic or other molecular-based conditions that require lifelong treatment. Molecular biomarker tests and genome-sequencing technologies offer the ability to identify conditions early in childhood to facilitate treatment that may avert significant morbidity and mortality in the long term. It is not always clear, however, how to position these tests in an assessment pathway. Chen et al.’s study examined the incremental cost-effectiveness of identifying phenylketonuria, a genetic disorder, via newborn screening with a genetic test, compared with a symptom-based clinical diagnosis over the lifetime from a societal payer perspective . Their mThis collection touches on critically important concepts in understanding value in child health and should inspire researchers and decision makers alike to think more broadly when considering how to value interventions and services that benefit children and their families.


Growing numbers of people with multimorbidity have a co-occurring mental health condition such as depression. Co-occurring depression is associated with poor patient outcomes and increased healthcare costs including unplanned use of secondary healthcare which may be avoidable.To summarise the current evidence on the association between depression and unplanned secondary healthcare use among patients with multimorbidity.We conducted a systematic review by searching MEDLINE, EMBASE, PsychINFO, Web of Science, CINAHL, and Cochrane Library from January 2000 to March 2021. We included studies on adults with depression and at least one other physical long-term condition that examined risk of emergency hospital admissions as a primary outcome, alongside emergency department visits or emergency readmissions. Studies were assessed for risk of bias using The National Institute of Health National Heart, Lung, and Blood Institute quality assessment tool. Relevant data were extracted from studies and a narrative synthesis of findings produced.Twenty observational studies were included in the review. Depression was significantly associated with different outcomes of unplanned secondary healthcare use, across various comorbidities. Among the studies examining these outcomes, depression predicted emergency department visits in 7 out of 9 studies; emergency hospital admissions in 19 out of 20 studies; and emergency readmissions in 4 out of 4 studies. This effect increased with greater severity of depression. Other predictors of unplanned secondary care reported include increased age, being female, and presence of greater numbers of comorbidities.Depression predicted increased risk of unplanned secondary healthcare use in individuals with multimorbidity. The literature indicates a research gap in identifying and understanding the impact of complex multimorbidity combinations, and other patient characteristics on unplanned care in patients with depression. Findings indicate the need to improve planned care for patients with moderate-to-severe depression. We suggest regular reviews of care plans, depression severity monitoring and assessment of hospital admission risk in primary care settings. Multimorbidity, the co-occurrence of two or more chronic conditions , is incrIndividuals with multimorbidity have complex needs requiring long-term management and treatment of multiple conditions across multiple healthcare settings. Multimorbidity, in general, is associated with increased health service utilisation across primary care, planned secondary care, urgent care and dental care –13 and wSpecifically, the use of unplanned secondary healthcare may be considered as a proxy for inadequate or unsuitable planned care for individuals with multimorbidity , 17 and,A previous systematic review and meta-analysis  conducteThe aim of this review is to summarise the current evidence on the association between depression and unplanned secondary healthcare use among patients with multimorbidity. To our knowledge, the literature is limited on specifically depression-related multimorbidity clusters, namely different combinations of comorbidities, or specific patient characteristics and the subsequent effect on unplanned secondary healthcare use. Therefore, this review also aims to explore the effect of the types of comorbidities and if available, different clusters of comorbidities, and sociodemographic predictors of unplanned secondary healthcare among patients with both multimorbidity and depression.A review of the current literature was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines . An initStudies were eligible if they included adults aged 18+ with at least one other long-term condition in addition to depression and considered emergency hospital admissions as one of their main outcomes. Observational studies and randomised controlled studies comparing individuals with and without depression were considered for inclusion. Only studies using a standardised and validated measure of identifying depression in patients and reporting quantified measures of association such as risk ratios, odds ratios, or hazard ratios were included. Studies were excluded if participants were already in an emergency department or a current inpatient at the time of recruitment to the study. This is because the focus of our outcome is to determine the risk of an individual coming in to use the emergency department, including those with readmission risk, not for patients already found there. No filter restrictions were applied except for publication date.Searches in Medline, Embase, PsycINFO, Web of Science, CINAHL and Cochrane Library were performed in March 2021, for records since January 2000. Reference lists of relevant studies were hand-searched for further relevant publications. A combination of search terms was used to identify relevant articles, such as “multiple long-term condition/comorbid”, “depression”, and “emergency care”. The search strategy was developed with the assistance of a specialist librarian at Imperial College London. Search terms and syntax were adapted for each individual database .Records captured through electronic database searching and hand-searching were de-duplicated. Two reviewers (MC and MO) independently screened the titles and abstracts of the remaining records for inclusion based on eligibility criteria. The same two reviewers independently reviewed the full text of eligible articles. Discrepancies were resolved through discussion with the other authors.A data extraction form was developed and used to capture key data from records relevant to the review aims. Data extracted included study characteristics, sociodemographic and clinical characteristics associated with emergency hospital admissions and emergency department visits as per the study findings, and measure of risk of unplanned secondary healthcare use in respective study populations. Primary data extraction was performed by MC and was reviewed by MO.Following full-text screening of relevant articles, two reviewers (MC & MO) independently appraised the included studies for the risk of bias by using the National Heart, Lung, and Blood Institute (NHBLI) quality assessment tool for observational and cross-sectional studies  see S3 . This toThe main outcome of interest was the risk of emergency hospital admission, and secondary to that emergency department visits and/or emergency hospital readmissions as measures of unplanned secondary healthcare use. The rationale for this is that patients who undergo emergency hospital admission will have likely passed through the emergency department, thus capturing the outcome more likely to indicate severe patient condition.Evidence tables were produced to include relevant data on the study characteristics, clinical and sociodemographic predictors, and key findings. Certainty of outcome measures  reported were assessed prior to synthesis based on 95% confidence intervals and statistical significance threshold of p<0.05. A narrative synthesis of the data was conducted to summarise key findings by grouping studies with similar outcomes, clinical and sociodemographic predictors of unplanned secondary healthcare, as available in included studies. Data collected from the studies were not suitable for meta-analysis due to the heterogeneity of the outcome measures across the studies.Database searches identified 11,228 records, with three identified through hand-searching. After removing 2,874 duplicates, 8,384 titles and abstracts were screened, of which 8,352 did not meet the eligibility criteria. Full texts were obtained for thirty-two articles of which a further 12 articles were excluded see .17 studies were based in high-income settings including USA (n = 12) –37, 39, All studies clearly defined their objective and study population; recruited participants from the same population with uniform eligibility criteria applied; clearly defined and implemented both exposure and outcome measures consistently across study participants; measured and adjusted for key potential confounders see . All butVarious methods and tools were used to ascertain depression status in studies’ samples. This included depression scales (n = 6) such as the Hospital Anxiety and Depression Scale (HADS) , 40, 41 Index comorbidities refer to the main condition of focus in each study, given that participants had at least two simultaneous chronic conditions. A range of comorbidities were observed across the included studies ; the mosAll 20 studies considered emergency hospital admissions as an outcome, while nine also looked at emergency department (ED) visits , 33–36 aSeven out of nine studies that studied ED visits as an outcome demonstrated that depression predicted increased visits e.g., OR = 2.45 in cancer patients  and HR =Depression predicted emergency hospital admissions in 19 out of 20 studies; some studies demonstrated that this positive association was proportional to increasing increments of depression severity , 40, 41.The reasons for emergency hospital admissions were not necessarily mentioned across all studies, although some distinguished admissions as being specific to the index comorbidities studied. For example, Bhatt and colleagues  report hIn a few of the studies investigating both emergency hospital admissions and ED visits, depression predicted one outcome but not the other –27. For Four studies investigated the risk of readmission in patients with various multimorbidity profiles and depression, which all reported a statistically significant greater risk for readmission in those with depression , 39, 40.Although most studies adjusted results for sociodemographic covariates, only one study  reportedNo studies analysed the risk of any type of unplanned secondary healthcare use by multimorbidity disease combinations or clusters; index conditions were individually studied without further exploration of the cumulative effects of multiple conditions in the same individuals. Though not a risk estimate, one study showed that the number of long-term conditions were positively correlated with a greater percentage of patients using unplanned secondary care . Those wThis review found that presence of depression increases the likelihood of emergency hospital admissions and readmissions in patients with multimorbidity. This association holds across a range of long-term conditions characterising multimorbidity in various countries, settings and samples. Depression also predicted increased ED visits in most of the studies reporting on this outcome. Moreover, the greater the severity of depression, the greater the risk of emergency hospital admissions and ED visits.Patients with co-occurring depression with cancers , COPD 2, and astThis review captures a broad scope of multimorbidity, inclusive of any chronic comorbidity, in an area of growing importance and contributes up-to-date evidence that adds to a previous systematic review. We focussed on unplanned secondary healthcare use instead of all types of unplanned care, such as urgent GP home visits, use of walk-in centres and minor injuries units, which allowed for more uniform comparison and synthesis of study findings. We conducted an extensive search of six electronic databases with a comprehensive search strategy designed to capture a broad range of multimorbidity studies without limiting comorbidity, publication language, sample size or follow-up duration. Our review additionally highlights the effect of incremental depression severity on the studied association, beyond establishing a simple association. Through excluding studies that recruited individuals from the emergency department itself or during an active inpatient period, we aimed to minimise confounding as much as possible.However, some limitations exist. Firstly, the range of different conditions included in the review may have inherently different effects to each other on the studied outcome, based on whether some conditions are more prone to adverse events requiring greater emergency care episodes than others. For example, conditions like COPD or heart failure have varying severity levels which are highly sensitive to exacerbations or adverse events, compared to other conditions like hypertension or diabetes where effects may be relatively slower to accumulate. Secondly, the use of depression scales in some studies only measures status and severity of depression cross-sectionally, which does not capture the onset and type of depression, namely episodic versus recurrent. We are not able to deduce what the temporal effect of depression is based on duration of illness before and during study periods to consider the bidirectional relationship with long-term conditions. Thirdly, sample sizes and follow-up periods varied across the studies which may explain some of the variation in the effect sizes. For example, two studies reporting diabetes-related emergency admissions conclude opposite findings which may be related to sample size and outcome duration of hypoglycaemic episodes due to different follow-up periods , 30. AlsThis review concurs with the overall positive association between depression and unplanned secondary healthcare found in the previous review on all types of unscheduled care for four selected comorbidities . The preWe found that incremental increases in depression severity scores were associated with an increase in the magnitude of risk of unplanned secondary care outcomes, particularly for COPD  and hearThe strong association between increased depression severity and increased unplanned secondary healthcare use suggests limitations in the adequacy of planned care and ongoing management of physical comorbidity in individuals with co-occurring depression. Particular attention must be paid to those with high depression scores to ensure the depression is optimally treated, to review clinical management plans, and utilise other resources such as integrated multidisciplinary teams to reduce avoidable unplanned care . DepressThis review suggests a gap in our understanding of how the risk of using unplanned secondary healthcare may vary with multimorbidity clusters i.e., the broader spectrum of disease combinations that make up patients’ multimorbidity. This is important to investigate in patients, particularly those with complex multimorbidity which is becoming more common, namely those with more than two conditions belonging to multiple body systems. Additionally, there is limited experimental evidence on whether interventions to treat depression in people with multimorbidity reduce unplanned care use. Nor is there sufficient evidence on whether depression interacts with certain patient characteristics to exacerbate the risk of emergency hospital admissions among individuals with multimorbidity. This gap requires investigating the stratified risk for different patient groups and their heterogenous multimorbidity profiles in future research. Furthermore, future research should seek to understand the reasons for increased emergency care in patients with depression, using mixed methods approaches.Concurrent depression is an important predictor of unplanned secondary healthcare use in individuals with multimorbidity, with this effect appearing to increase with greater severity of depression. More research is needed to identify multimorbidity clusters and other patient characteristics which may predict unplanned care use as well as the reasons for this effect in depression. Individuals with depression and multimorbidity should be proactively identified in the primary care setting and action taken (through depression severity and admission risk scoring tools) to identify those at particularly high risk of unplanned care use, to conduct regular primary care reviews, assessment, and treatment to reduce this risk.S1 Table(DOCX)Click here for additional data file.S2 TableSearch terms and combinations for Medline, Embase, PsychINFO, Web of Science, CENTRAL, CINAHL databases, performed in March 2021.(DOCX)Click here for additional data file.S3 Tablehttps://www.nhlbi.nih.gov/health-topics/study-quality-assessment-tools). Legend: Yes (Y); No (N); Other (Not Applicable (N/A); Not Reported (N/R); Cannot Determine (C/D)). Overall Rating: Good, Fair, Poor.Results using the National Heart, Lung, and Blood Institute (NHBLI) Study Quality Assessment Tools (Available at: (DOCX)Click here for additional data file.


The emergent spatial and temporal patterns of adaptation, observed as changes in cellular connectivity and tissue remodeling by cells, underpin neuroplasticity, muscle memory, immunological imprinting, and sentience itself, in diverse physiological systems from brain to bone. Connectomics addresses the direct connectivity of cells and cells’ adaptation to dynamic environments through manufacture of extracellular matrix, forming tissues and architectures comprising interacting organs and systems of organisms. There is imperative to understand the physical renderings of cellular experience throughout life, from the time of emergence, to growth, adaptation and aging-associated degeneration of tissues. Here we address this need through development of technological approaches that incorporate cross length scale (nm to m) structural data, acquired via multibeam scanning electron microscopy, with machine learning and information transfer using network modeling approaches. This pilot case study uses cutting edge imaging methods for nano- to meso-scale study of cellular inhabitants within human hip tissue resected during the normal course of hip replacement surgery. We discuss the technical approach and workflow and identify the resulting opportunities as well as pitfalls to avoid, delineating a path for cellular connectomics studies in diverse tissue/organ environments and their interactions within organisms and across species. Finally, we discuss the implications of the outlined approach for neuromechanics and the control of physical behavior and neuromuscular training.“Brainless” cells, the living constituents inhabiting all biological materials, exhibit remarkably  Cells of the human body populate their habitat through division, starting with two cells at conception and expanding to over 70 trillion cells over the course of a lifetime . Throughemergent properties , and other imaging modalities that enable high temporal albeit less spatial resolution (MRI). Rapid advances in the field seek to overcome this current hurdle. To address each of these points, workflows are needed to render and analyze vast amounts of imaging data from nano- to meso- length scales. The manuscript describes that process and sets a path forward.g., in the brain, as the connectome and the process of rendering, analyzing and understanding the connectome as connectomics  networks, e.ectomics . A recenectomics . Tested ectomics  and in oectomics , as wellectomics , these bHere, key enabling steps are described for quantifying relationships and connectivity between cells in different disease states. Specifically, we test machine learning algorithms with cellular network maps of the human hip to elucidate the role of cell networks in organ and organism (patho)physiology throughout life . This apHuman tissue samples from the femoral neck and head of patients undergoing total hip replacement were obtained with Institutional Review Board Approval (Cleveland Clinic IRB12-335). Samples were prepared for electron microscopy (EM) using a published protocol  developeSamples were first etched (to expose cells just below the block face surface) and imaged using multi-beam Scanning Electron Microscopy, to achieve nano- to meso-scale renderings of cellular inhabitants (mainly osteocytes with some red blood cells visible in resulting images). Based on this protocol, carbon coating provided sufficient contrast to visualize osteocytes exposed by chemical etching on the surface of the sample block. Although not used for the current study, sequential layers of cellular networks could be revealed by reiterating the etching and imaging steps, resulting in a volume of tissue with fully rendered three dimensional (3D) cellular network.Three datasets were acquired using three generations of multibeam Scanning Electron Microscopes (mSEM) to image three different samples, starting with a 61 beam prototype at 12 nm pixel size and ending with a state-of-the-art commercial system (Zeiss mSEM 505) .pyknotic cells . As a surrogate identification factor (classifier for machine learning implementation), osteocytes with less than three visible processes were identified visually and manually pinned as pyknotic, indicated by a red pin. Osteocytes with more than three visible processes were identified visually and manually pinned as viable, indicated by a green pin. The manual process took several weeks for the 1st generation dataset  machine learning algorithm , using t2 tissue and over 50,000 images (0.08 terabyte), with manual identification of 629 osteocytes taking several weeks’ time. To enable automated, rapid detection of osteocytes and in consideration of the increasing size and complexity of the datasets enabled through advances in the mSEM instrument and parallel computational advances (second and third generations) over the past decade , we applied a machine learning algorithm to detect osteocytes based on the You Only Look Once (YOLO) convolutional neural network described originally by In summary, we acquired three datasets rendering osteocyte networks in tissues of the femoral head. The different datasets include data from different samples imaged using new generations of mSEM and associated increasing computational capacity. The first data set included 5.69 mm2 tissue and over 7,000,000 images (10.98 TB), identifying a total of 206,180 osteocytes in 100 h on a graphics processing unit  compared to the manual pinning method of our previously published work  under vacuum . FollowiManual marking of landmarks was described in a previous paper  and the described further for the layperson inThe “You Only Look Once” (YOLO) neural network automated object detection algorithm (6 examples through variation by rotation, scale and contrast (Initially, YOLO was trained for automated osteocyte detection, using 629 annotated cells, which were further augmented to 10contrast . Unseen contrast  to obtaiThe original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.The studies involving human participants were reviewed and approved by the Cleveland Clinic Institutional Review Board.MK and DZ conceived the work. The specimens were collected and prepared for imaging by MK with assistance from the Cleveland Clinic and Zeiss teams. Image acquisition was carried out by the Zeiss Team, led by DZ and MK in the Demonstration Labs at Carl Zeiss Microscopy GmbH in Oberkochen. The machine learning algorithm was developed and tested by AS and CW. The manuscript was written by MK and was revised and approved of by all coauthors. All authors contributed to the article and approved the submitted version.Zeiss provided in kind support for this project, which is of a fundamental and translational nature. AS and CW were employed by Zeiss AG. DZ was employed by Carl Zeiss MultiSEM GmbH. The remaining author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Using data from the 2006, 2012, and 2018 waves of the Health and Retirement Study, we estimated effects of co-occurrence of diabetes mellitus (DM) and complete tooth loss (CTL), both self-reported, on cognitive function among 10,816 adults age 50+. Cognitive function was measured using a shortened version of the Telephone Interview for Cognitive Status. Results from the fixed effects linear regression model show that in comparison to those with neither condition, adults having both DM and CTL had the worst cognitive function , followed by having CTL alone , and having DM alone . Our study suggests that CTL is a stronger risk factor for lower cognitive function than DM, and the co-occurrence of DM and CTL poses additive risk. Further research is needed to investigate the pathway from DM and CTL to poor cognition.


Viruses are capable of hijacking normal cellular functions, and understanding the role of BAs and BA receptors, components of the enterohepatic system, is valuable for expanding our knowledge on the mechanisms of norovirus and hepatitis B and C virus replication.Laboratory cultivation of viruses is critical for determining requirements for viral replication, developing detection methods, identifying drug targets, and developing antivirals. Several viruses have a history of recalcitrance towards robust replication in laboratory cell lines, including human noroviruses and hepatitis B and C viruses. These viruses have tropism for tissue components of the enterohepatic circulation system: the intestine and liver, respectively. The purpose of this review is to discuss how key enterohepatic signaling molecules, bile acids (BAs), and BA receptors are involved in the replication of these viruses and how manipulation of these factors was useful in the development and/or optimization of culture systems for these viruses. BAs have replication-promoting activities through several key mechanisms: (1) affecting cellular uptake, membrane lipid composition, and endocytic acidification; (2) directly interacting with viral capsids to influence binding to cells; and (3) modulating the innate immune response. Additionally, expression of the Na Viruses are uniquely fascinating and challenging to study because they are obligate intracellular pathogens that often evolve a tropism not only for their specific hosts but for specific tissue and cell types. Molecular and physical techniques or animal models can be used to identify, detect, and study viral pathogenesis. However, cell culture is ideal for larger scale propagation of viruses, detailed studies of the mechanisms of viral replication, and studying the efficacy of antibodies and antivirals. Cell lines are often easy to genetically manipulate—a property that can be used to determine host factors required for infection. Several important human viruses, including hepatitis viruses and caliciviruses, remain difficult to cultivate despite advances in cell culture technology. Understanding cellular tropism and host factors was critical for the development of efficient systems to propagate these viruses. For the cultivation of hepatitis B virus (HBV) and human noroviruses (HuNoVs) that infect the liver and the intestine, respectively, bile components including bile acids (BAs) and BA receptors are now known to be key for successful replication ,2. BAs a+-taurocholate cotransporting polypeptide (NTCP) for recycling to the liver up to 4–12 times per day with the recycling pool containing primarily CA, CDCA, DCA, and trace amounts of LCA  had earlier established culture systems, defined cellular receptors and are often used as surrogates for HuNoV infection . D. D67]. DViruses are unique engineers that, through evolution, exploit host cellular functions for the most effective replication tools. Noroviruses and hepatitis viruses have utilized host enterohepatic circulation factors  to allow their entry and replication in different human cells  makes this a candidate to test as a therapeutic for multiple strains of HuNoV infection. Inhibitors such as MyrB are being investigated for the potential to target NTCP for the development of HBV therapeutics . These p


Traditional proximate analysis measures  have little value for estimating the actual nutritional and economic value of feed ingredients fed to swine, yet they unfortunately continue to be commonly used in research studies, feed label regulations, and commodity-trading contract guarantees. Increasing energy and nutritional efficiency while simultaneously reducing negative environmental impacts of pork production requires the adoption of precision nutrition practices in global pork production systems. Precision swine nutrition can only be achieved by using more accurate and comprehensive methods and measures to determine the true nutri-physiological value of feed ingredients used in swine diets. There are several characteristics in some types of feed ingredients that are detrimental to swine health and performance, but they are seldom measured and deserve greater consideration. Likewise, there are also several value-added benefits provided by some functional feed ingredients that are not given enough consideration when formulating swine diets. The purpose of this review is to provide a holistic assessment of the benefits and limitations of existing chemical analysis methods and measures used to evaluate feed ingredients for swine and explore the benefits of using additional measurements to more accurately assess their true nutri-physiological value.Many types of feed ingredients are used to provide energy and nutrients to meet the nutritional requirements of swine. However, the analytical methods and measures used to determine the true nutritional and physiological  value of feed ingredients affect the accuracy of predicting and achieving desired animal responses. Some chemical characteristics of feed ingredients are detrimental to pig health and performance, while functional components in other ingredients provide beneficial health effects beyond their nutritional value when included in complete swine diets. Traditional analytical procedures and measures are useful for determining energy and nutrient digestibility of feed ingredients, but do not adequately assess their true physiological or biological value. Prediction equations, along with ex vivo and in vitro methods, provide some benefits for assessing the nutri-physiological value of feed ingredients compared with in vivo determinations, but they also have some limitations. Determining the digestion kinetics of the different chemical components of feed ingredients, understanding how circadian rhythms affect feeding behavior and the gastrointestinal microbiome of pigs, and accounting for the functional properties of many feed ingredients in diet formulation are the emerging innovations that will facilitate improvements in precision swine nutrition and environmental sustainability in global pork-production systems. Multifaceted approach is emerging for selecting and sourcing feed ingredients in the global feed industry to achieve biosecure, econutritional, precision swine-feeding programs without the use of growth-promoting antibiotics. Historically, feed ingredients have been primarily selected based on availability and cost, as well as crude protein (CP), crude fat, and crude fiber content ,2. HowevAll of these important criteria for selecting and sourcing feed ingredients must also be merged with precision swine nutrition practices, which are a key component of precision livestock farming. Precision livestock farming  is a rapidly emerging, innovative, systems approach that is being implemented to improve productivity and sustainability of food animal production to meet the increasing need for safe and affordable food to feed a growing global population . As precAlthough the determination of the true nutri-physiological value of feed ingredients and complete feeds has greatly improved over time, significant challenges still exist . InitialFortunately, in vivo methodologies were developed to determine energy and nutrient digestibility of feed ingredients and complete feeds for swine, which was the beginning of estimating the actual nutritional contributions of feed ingredients from their chemical composition . As new Despite these nutritional advances, some limitations of using conventional evaluation and analytical methods remain for assessing the utilization of dietary energy , AAs 3030, and pThere are many challenges for determining the true nutri-physiological value of feed ingredients, including the development and standardization of accurate methods for estimating bioavailability of nutrients that are additive in mixtures of feed ingredients , and chaFeed ingredients are routinely evaluated both qualitatively and quantitatively to determine their perceived and real nutritional and economic value. Physical characteristics such as color, odor, and texture are subjective qualitative assessments that can be useful for initial screening of general acceptability of ingredient quality, but they are of limited value for assessing nutrient composition, digestibility, and presence of antinutritional factors. As a result, various chemical analyses have been developed and used to provide quantitative determinations of nutrient content, contaminants, and toxic compounds. However, results from chemical analyses vary among laboratories using the same procedure, as well as among different procedures used for measuring the same analyte . In addiw) may be even more important for understanding the nutri-physiological value of feed ingredients, and it is a commonly used measurement in human foods [Many animal nutritionists and feed manufacturers give little consideration to the moisture content of feed ingredients other than ensuring that it is low enough to avoid spoilage during long-term storage and to optimize pellet manufacturing conditions. Although measuring the moisture content of feed ingredients is generally considered to be simple and accurate, it is actually more difficult and complex than many nutritionists realize, and requires greater scrutiny because of its association with nutrient preservation and feeding value of ingredients. Moisture content can be determined using 35 different direct or indirect methods described by the Association of Official Analytical Collaboration (AOAC) International. Direct methods  are generally considered to be the most reliable measures but are more labor-intensive and time-consuming than indirect methods . Furthermore, calculations of the percentage of moisture in a feed sample are often assumed to be interchangeable between reporting moisture on a wet or a dry basis. However, they are not interchangeable because when calculating moisture content on a wet basis, the amount of water is divided by the total weight of the sample (solids and moisture), while on a dry basis, the amount of water is divided by dry weight . Therefan foods ,44 and pan foods , but notw is useful for predicting safety and stability of food and feed products because microbial growth is determined by water activity and not by water content, as commonly believed. Water activity is useful for predicting the types of microorganisms that may cause spoilage [Water activity is a measure of the energy state of water in food or feed and its potential to act as a solvent, participate in biochemical reactions, and support growth of microorganisms. The measurement of water activity was developed because moisture content of a food did not adequately represent the fluctuations in microbial growth that were commonly observed. Therefore, aspoilage ,45 and mspoilage . Specifispoilage , nonenzyspoilage ,48, enzyspoilage ,50, and spoilage ,52. In aspoilage  and pellspoilage  are direw of 0.61 is the minimum value that allows fungal growth [w, where milling to a fine particle size appears to reduce water activity  of feeds that is utilized for maintenance and productive functions of animals . The GE Systems used for estimating energy requirements and utilization in feed ingredients for swine have evolved from DE to ME to net energy (NE) determinations. Net energy is the actual energy remaining for maintenance and productive purposes after accounting for additional energy losses from heat produced (heat increment) during ingestion, digestion, and physical activity beyond those attributed to ME . The NE Even though NE systems are more accurate than ME and DE systems, an inherent assumption is that the energy used for body protein and lipid gain, as well as body maintenance functions, is not different between pigs with different rates and composition of body weight gain . This asSeveral reviews have described the comparative advantages and disadvantages of using various methodologies for determining NE in feed ingredients ,65,66,67The ultimate goal of any energy system is to accurately rank feed ingredients based on their actual energy contribution for predicting pig performance. Because increasing amounts of high-fiber feed ingredients are being used in swine diets globally, use of the NE system is essential for avoiding overestimation of utilizable energy from these sources. In addition, use of the NE system minimizes the potential for overestimating energy utilization when formulating low-CP diets, which are becoming popular to minimize postweaning diarrhea and nitrogen excretion in manure for improved environmental sustainability. Because energy intake is the most limiting factor for protein deposition in young pigs, protein synthesis is an energetically expensive process, and energetic efficiencies to produce ATP or lipid synthesis are different , direct Carbohydrates are the primary chemical constituents of plant-based feed ingredients and serve as the primary energy source in swine diets. In addition, lactose and chitin are animal-derived carbohydrates that are present in a few feed ingredients fed to swine. Lactose is the main carbohydrate in milk for young mammals and serves as an energy source after it is hydrolyzed by lactase to glucose and galactose in the small intestine . The addPlant-based carbohydrates vary substantially in their monomeric composition, types of linkages, and extent of polymerization . As a reBecause of the diversity of physicochemical characteristics of numerous DF components, it has been challenging to relate traditional chemical measurements for DF to specific physiological effects. The analytical methods most commonly used for determining DF content of feed ingredients and complete feeds include crude fiber, van Soest methods , and total dietary fiber (TDF = soluble + insoluble dietary fiber) methods . The cruDietary fiber comprises many carbohydrate components that have several physicochemical properties related to physiological responses. McRorie and Fahey  developeThere is increasing interest and evidence indicating that the time of day when DF fermentation occurs is an important determinant of its utilization. Circadian rhythms affect eating behaviors, gut microbiome, nutrient utilization, and health of animals . Most phEscherichia coli [The physicochemical properties of DF can have detrimental or beneficial effects in swine diets. Numerous studies have shown that high-fiber diets generally decrease nutrient digestibility and increase endogenous nutrient loss, leading to a detrimental effect on energy metabolism and growth performance of pigs . Viscosihia coli ,102,103.hia coli ,105. Furhia coli . Rapidlyhia coli ,90. Howehia coli . AdditioFeeding high-fiber diets to sows provides several physiological and reproductive benefits . Limit-fLipids contain 2.25 times more energy than carbohydrates, but crude fat  content is poorly associated with ME and NE content of grains ,116, as One of the greatest challenges of capturing full nutritional and economic value of any feed ingredient is to use accurate energy values for specific sources used in swine diet formulations. Feed fats and oils contribute a substantial proportion of energy to swine diets, but like other feed ingredients, the chemical composition and DE, ME, and NE content is highly variable within and among sources . For exaFortunately, there continues to be considerable interest in developing and using robust equations to accurately estimate DE, ME, and NE content of lipids based on chemical composition . HoweverThere may be several reasons for the lack of accurate predictions of DE content observed in these studies. First, the equations are likely to be too simplistic and do not include other important predictive variables that may improve their accuracy and potential use. Second, if prediction equations are used to estimate energy content of fats and oils, they should be derived from experiments that have directly determined DE, ME, or NE content of those specific types of lipids, and not from studies that evaluated other types of ingredients or different lipid sources. Third, energy prediction equations derived from various lipids with diverse fatty acid profiles are assumed to provide reasonably accurate predictions when compared with energy values directly determined from in vivo experiments. However, results from Kellner and Patience  and KerrAlthough prediction equations may reasonably estimate DE, ME, or NE content of lipid sources in an experiment from which they were derived , they maFeed fats and oils, as well as the lipid fraction in other feed ingredients such as corn DDGS , are freIn the Weende analysis system, CP is defined as the nitrogen-containing fraction of a feed ingredient ,27. CrudBecause protein is the second most expensive component of swine diets, and represents a significant portion of the total diet, accurate estimation of AA content in feed ingredients is needed to make real-time adjustments in databases used for precision swine-diet formulation . In geneMore importantly, nutritionists need methods to dynamically estimate digestible and bioavailable AA content of feed ingredients used in precision diet formulation. Several reviews have been published describing the advantages and disadvantages of using various in vivo experimental techniques and procedures for determining AA digestibility in feed ingredients for swine ,33,172. When feed ingredients are subjected to heat treatment during processing, the epsilon amino group of free and protein-bound lysine may react with reducing sugars to create several types of Maillard products ,177. In 2 = 0.72 to 0.88) between total concentrations of all indispensable AAs and AID of AAs, but correlations differed between ingredients due to the negative effect of NDF content on AID in some ingredients. Although the prediction accuracy of all new SID equations developed was improved compared with original versions, they require validation before being used in practical swine diet formulations [Development and use of prediction equations to estimate SID of AAs in feed ingredients based on chemical composition has also been considered as an alternative approach to overcome the challenges of using the various in vitro methods. Zeng et al.  conducteulations . Other aulations .Perhaps the most promising approach for predicting biological responses from chemical composition of feed ingredients involves using LC-MS, advanced proteomic, and bioinformatic approaches . The nutProtein oxidation of feed ingredients is an emerging area of concern because of its potential contributions toward exacerbating oxidative stress in animals . ProteinPhosphorus is the third most expensive component of swine diets and requires special consideration for optimizing its utilization in swine diets due to its potential adverse effects on the environment. Calcium and phosphorus are the most abundant minerals in animals, and about 90% of Ca and 80% of P in the body is concentrated in the skeleton . Feed inHistorically, P bioavailability estimates were used for assessing the proportion of total P that was digested and absorbed by the animal ,211. HowTo overcome this problem, methodologies to determine apparent (ATTD), standardized (STTD), and true total tract digestibility (TTTD) of Ca and P have been used to more accurately express their utilization in various feed ingredients for swine ,33. UnliTo optimize P utilization when feeding diets containing plant-based ingredients containing variable but relatively high concentrations of phytate to swine, exogenous phytase enzymes must be added to increase the proportion of dietary P used by the animal, reduce P excretion in manure, and minimize the antinutritional effects of phytate on digestibility of other nutrients ,217,218.The use of in vivo models is generally considered to be more accurate for assessing the nutri-physiological properties of feed ingredients compared with in vitro and ex vivo methodologies. However, in vivo models have the disadvantages of being subjected to confounding factors such as uncontrolled environmental conditions, differences in microbiome and immune status of animals, and substantial variability in individual pig responses within the same dietary treatment. Because of these confounding factors, use of in vitro and ex vivo methodologies provide several advantages for specific applications when evaluating feed ingredients. First, they provide a faster and less-expensive means for dynamically estimating the nutri-physiological value of feed ingredients compared with expensive and cumbersome in vivo methods. In fact, some nutrition researchers have recognized that holistic and accurate evaluation of the nutritional value of feedstuffs often requires separate assessment of specific components of digestive processes and their end products at the specific sites where the nutrient digestion and absorption occurs along the gastrointestinal tract . FurtherSeveral in vitro methods have been developed and used for estimating nutrient digestibility in feed ingredients during the past several decades . AlthougNear-infrared reflectance spectroscopy is a rapid, physically nondestructive, and inexpensive technique that is rapidly being adopted in the global feed industry to accurately estimate nutrient content of feed ingredients. The principle of this technique involves measuring the reflectance spectrum in the near-infrared wavelength region of the sample and comparing these data with reference spectra of known samples to provide a quantitative estimate of analytes of interest. Calibration of NIRS equipment is based on using nutrient-composition data derived from standard chemical analysis procedures and various statistical techniques. The accuracy and precision of NIRS estimates are dependent on the robustness and size of the data set used to create the calibration model. Near-infrared reflectance spectroscopy calibrations have been developed for determining proximate analysis components ,229,230,(1)Rapid scanning of the samples (less than 1 min).(2)Only a small amount of sample is needed for analysis.(3)Low cost because no chemical reagents are needed and a single operator can analyze a large number of samples in a short period of time.(4)Results are highly reproducible.(5)Multiple analytes can be determined in one operation.(6)Minimal (drying and grinding) or no sample preparation is needed.(7)Equipment can easily be used in different environments .(8)High accessibility for online data capture and storage.(9)Some optical probes allow analyzing samples in situ.(10)Equipment is portable.Advantages(1)It is a secondary method that requires the use of data derived from chemical analysis or in vivo studies as reference values.(2)A large number of samples with variable composition and data with large variation is required for accurate and robust calibrations.(3)Highly trained personnel are required for calibration and validation of the results.(4)Continuous maintenance and updating of the calibration database is required.(5)Changes in chemical structure of nutrients that occur during the digestion process cannot be predicted using NIRS technology.(6)High initial cost for purchasing NIRS instruments.DisadvantagesClosed filtration or multi-enzymatic methods have been developed to mimic part or all of the in vivo gastrointestinal tract digestion process and are used to estimate the dry matter and nutrient digestibility of various ingredients by using single or multiple enzymes and collecting undigested residues. Świȩch  summariz2 = 0.92; [Historically, various two-step multienzymatic incubation methods, including pepsin–pancreatin ,244,245, = 0.92; ). Unfort = 0.92; .2 = 0.91) between in vitro P digestibility and in vivo STTD of P among 13 sources of rendered animal protein meals for swine. These results suggest that the two-step in vitro P digestibility assay is capable of good prediction of in vivo P digestibility of animal protein byproducts fed to swine.The two-step enzymatic method has also been applied to predict in vitro STTD of phosphorus (P) in poultry studies ,258,259,The three-step in vitro system is designed to estimate nutrient digestibility of the entire gastrointestinal tract of pigs, is highly repeatable, and has been extensively evaluated in several independent studies. The first two steps of this method involve consecutive incubations of feed ingredients with various enzymes that mimic digestion in the stomach, small intestine, and large intestine ,251,252.Despite these encouraging results, none of these in vitro methods completely simulate the complex biochemical and physiological events that occur during the in vivo processes of energy  and nutrIn general, data derived from these in vitro procedures do not match in vivo determinations for nutrient digestibility of feed ingredients. However, two- and three-step in vitro assays can provide relatively rapid and cost-effective analysis, and serve as reasonable initial screening methods for assessing the magnitude of digestibility and fermentability differences among different sources of ingredients . FurtherThe Ussing chamber system was first developed and introduced in the 1950s and is used for diffusion- or electrophysiology-based measurements . Ussing The major advantages of using Ussing chambers are to study regional  permeability and barrier function. However, the main technical challenge of conducting experiments with Ussing chambers is the limited amount of time (up to 2.5 h) of maintaining viable intestinal tissue, which is not enough for the investigation of the reversible disruption of tight junctions and extensive metabolism determinations ,279. TheUse of intestinal cell cultures offers many potential benefits for studying specific and complex mechanisms involving the chemical and nutritional composition of feed ingredients associated with physiological responses of pigs. Historically, two-dimensional intestinal cell line models, such as Caco-2 , HT-29 ,296, HCTEnteroids are three-dimensional structures that originate from embryonic stem cells, induced pluripotent cells, or adult stem cells from intestinal tissue, and are grown in cell culture. Unlike two-dimensional cell lines, enteroids have all of the differentiated cells found in the small intestine, including lumen, villi and crypts of enterocytes, enteroendocrine cells, goblet cells, tuft cells, Paneth cells, and stem cells . TherefoCompared with other in vitro models, use of enteroids provides the opportunity to study the effects of diet and nutrients on intestinal growth and development, ion and nutrient transport, secretory and absorptive functions, intestinal barrier, cell differentiation, intestinal disease, gene expression, and other responses of interest in the gastrointestinal tract ,304,305.Knowledge of and interest in using enteroid-based technologies continues to increase relative to their many advantages for animal nutrition research. Although limited studies have used enteroid models in swine-nutrition research, enteroids represent a promising ex vivo technology that could greatly enhance our knowledge of nutri-physiological characteristics of feed ingredients for swine in the future.Traditional feed ingredient evaluation models have described nutrient digestion, absorption, and metabolism as sequential events in which nutrients are made available for growth in a way that is quantified by the total amount of nutrient digested and absorbed in the gastrointestinal tract . For exaThe inefficiency of nitrogen utilization may be attributed to the concept that protein deposition is considered as an “all or nothing” event, in which all AAs must be present at the exact moment of protein synthesis or they will not be utilized . AssuminThe rate and amount of AA disappearance along the small intestine of pigs varies among feed ingredients , where ADigestion and absorption of simple carbohydrates, such as glucose and sucrose, require less time than that of more complex carbohydrates such as corn starch . DiffereDigestion and absorption kinetics of starch and protein rich ingredients have been the most studied and documented relative to pig growth performance and health, but these characteristics are less defined and described for high-fiber feed ingredients. There are, however, clear differences among feed ingredients in the site and rate of degradation of fiber in the gastrointestinal tract of pigs . The traFor example, in pigs consuming diets containing corn DDGS, which contains high concentrations of insoluble fiber, about 20.6% of the fiber disappears in the small intestine within about 8–12 h . TherefoDetermination of kinetics of DF degradation using three-step in vitro digestibility systems has begun to provide insight regarding the rate and type of DF degradation in common high-fiber feed ingredients. In vitro, fiber degradation may occur by fiber solubilization in the stomach (step 1) and small intestine (step 2) during the pepsin and pancreatin digestion phase, in which pepsin degradation requires 2 h duration, while pancreatin degradation lasts for 4 h. Additional DF disappearance from fermentation is measured using fecal inoculum in step 3. Studies have also been conducted to evaluate the use of nonstarch polysaccharide (NSP) degrading enzymes, which have shown increased disappearance of DF in wheat middlings and corn DDGS in the small intestinal phase of digestion . HoweverSeveral studies have been conducted to evaluate the application of using kinetics of protein, starch, and fiber degradation of feed ingredients in diet formulations to improve growth performance and gut health of nursery pigs ,326,327.When starch is degraded and absorbed in the small intestine, it serves as the primary source of energy in diets for pigs, and it is also a potential source of energy for microbes in the large intestine. Providing energy and nutrients to beneficial microflora in the large intestine is considered necessary for decreasing postweaning diarrhea and maintaining gut health in pigs. Consequently, understanding the kinetics of starch digestion may also be beneficial for enhancing growth performance of pigs. The kinetics of starch digestion has been measured by the amylase diffusion rate, and it appears that formulating diets with sources of starch that have high intrinsic digestibility increases growth performance of nursery pigs . This enIn summary, the rate and sites of protein, starch, and DF degradation along the gastrointestinal tract differs among feed ingredient sources, and the kinetics of degradation can be measured using in vitro and in vivo models. Although the use of protein, starch, and DF degradation kinetics in practical diet formulation is still at the nascent stage, results from using this approach are promising for improving pig growth performance and health. Many of the current nutrient digestion kinetics models have only been applied to evaluating individual feed ingredients, and do not account for the significant interactions between protein and DF, as well as with other dietary nutrients in complete feeds. Characterizing these interactions will require more extensive modeling to accurately represent the complex and often nonlinear interactions among dietary components. Finally, databases that provide nutrient degradation kinetics data need to be developed to apply this new approach of feed ingredient evaluation in routine practical swine diet formulations.Functional foods have been defined as those that provide a health benefit beyond satisfying nutrient requirements in ways not anticipated by traditional nutrition science . ReviewsNumerous reviews have been published to describe the specific functional ingredients and nutrients along with their beneficial health effects when included in swine diets . The prep-coumaric, and caffeic acids [p-coumeric acid content appear to be found in the greatest concentrations in corn grain and bran compared with other grains and their respective bran fractions  should be avoided because they provide very limited useful information about the nutritional and physiological properties of feed ingredients for swine. Theuse of the net energy system, SID AA content, and STTD phosphorus content measures are encouraged for evaluating feed ingredients and formulating swine diets, but additional measures should be considered in feed ingredient databases, including: (1) water activity; (2) solubility, fermentability, viscosity, and prebiotic effects of dietary fiber; (3) lipid and protein oxidation measures; and (4) estimates of true bioavailability of amino acids, vitamins, and minerals. Furthermore, assessing the digestion kinetics of various dietary components  is a promising new approach that allows matching the various rates and quantities of nutrients digested in mixtures of diverse feed ingredients to optimize nutri-physiological responses in swine. Combining net energy and digestible nutrient prediction equations derived from meta-analyses of large and robust data sets with NIRS determinations of total nutrient content of feed ingredients may be the most practical and accurate approach for dynamic estimation of nutrient loading values for feed formulation. Use of two- and three-step in vitro digestibility systems, intestinal enteroids, and LC-MS platforms to characterize digestive and metabolic responses will be necessary for determining nutrient bioavailability of micronutrients and functional properties of various ingredients. Perhaps the biggest challenge for the future is to develop an effective systems biology approach for integrating large, complex data sets involving numerous nutritional and physiological response criteria so thathighly accurate, dynamic, mechanistic, and predictive mathematical models can be developed and implemented for enhancing precision swine nutrition.