Temporal trends and transmission dynamics of pre-treatment HIV-1 drug resistance within and between risk groups in Kenya, 1986-2020.

BACKGROUND: Evidence on the distribution of pre-treatment HIV-1 drug resistance (HIVDR) among risk groups is limited in Africa. We assessed the prevalence, trends and transmission dynamics of pre-treatment HIVDR within and between MSM, people who inject drugs (PWID), female sex workers (FSWs), heterosexuals (HETs) and perinatally infected children in Kenya. METHODS: HIV-1 partial pol sequences from antiretroviral-naive individuals collected from multiple sources between 1986 and 2020 were used. Pre-treatment reverse transcriptase inhibitor (RTI), PI and integrase inhibitor (INSTI) mutations were assessed using the Stanford HIVDR database. Phylogenetic methods were used to determine and date transmission clusters. RESULTS: Of 3567 sequences analysed, 550 (15.4%, 95% CI: 14.2-16.6) had at least one pre-treatment HIVDR mutation, which was most prevalent amongst children (41.3%), followed by PWID (31.0%), MSM (19.9%), FSWs (15.1%) and HETs (13.9%). Overall, pre-treatment HIVDR increased consistently, from 6.9% (before 2005) to 24.2% (2016-20). Among HETs, pre-treatment HIVDR increased from 6.6% (before 2005) to 20.2% (2011-15), but dropped to 6.5% (2016-20). Additionally, 32 clusters with shared pre-treatment HIVDR mutations were identified. The majority of clusters had R0 ≥ 1.0, indicating ongoing transmissions. The largest was a K103N cluster involving 16 MSM sequences sampled between 2010 and 2017, with an estimated time to the most recent common ancestor (tMRCA) of 2005 [95% higher posterior density (HPD), 2000-08], indicating propagation over 12 years. CONCLUSIONS: Compared to HETs, children and key populations had higher levels of pre-treatment HIVDR. Introduction of INSTIs after 2017 may have abrogated the increase in pre-treatment RTI mutations, albeit in the HET population only. Taken together, our findings underscore the need for targeted efforts towards equitable access to ART for children and key populations in Kenya.

Spatial clustering of livestock Anthrax events associated with agro-ecological zones in Kenya, 1957-2017.

BACKGROUND: Developing disease risk maps for priority endemic and episodic diseases is becoming increasingly important for more effective disease management, particularly in resource limited countries. For endemic and easily diagnosed diseases such as anthrax, using historical data to identify hotspots and start to define ecological risk factors of its occurrence is a plausible approach. Using 666 livestock anthrax events reported in Kenya over 60 years (1957-2017), we determined the temporal and spatial patterns of the disease as a step towards identifying and characterizing anthrax hotspots in the region. METHODS: Data were initially aggregated by administrative unit and later analyzed by agro-ecological zones (AEZ) to reveal anthrax spatio-temporal trends and patterns. Variations in the occurrence of anthrax events were estimated by fitting Poisson generalized linear mixed-effects models to the data with AEZs and calendar months as fixed effects and sub-counties as random effects. RESULTS: The country reported approximately 10 anthrax events annually, with the number increasing to as many as 50 annually by the year 2005. Spatial classification of the events in eight counties that reported the highest numbers revealed spatial clustering in certain administrative sub-counties, with 12% of the sub-counties responsible for over 30% of anthrax events, whereas 36% did not report any anthrax disease over the 60-year period. When segregated by AEZs, there was significantly greater risk of anthrax disease occurring in agro-alpine, high, and medium potential AEZs when compared to the agriculturally low potential arid and semi-arid AEZs of the country (p < 0.05). Interestingly, cattle were > 10 times more likely to be infected by B. anthracis than sheep, goats, or camels. There was lower risk of anthrax events in August (P = 0.034) and December (P = 0.061), months that follow long and short rain periods, respectively. CONCLUSION: Taken together, these findings suggest existence of certain geographic, ecological, and demographic risk factors that promote B. anthracis persistence and trasmission in the disease hotspots.

Integration of voluntary male medical circumcision for HIV prevention into norms of masculinity: findings from Kisumu, Kenya.

While it is clear that in many communities ideas about masculinity and circumcision are connected, it is still unclear how young Kenyan men in the former Nyanza province from the traditionally non-circumcising Luo people perceive voluntary medical male circumcision as connected to masculinity and the role of voluntary medical male circumcision in the transition from boyhood to manhood. The objective of this study was to explore norms of masculinity and the decision-making process among Luo young men to provide a better understanding of how circumcision and masculinity relate to cultural norms within this community. The methodology consisted of eight FGDs with male peer groups and 24 in-depth interviews to elicit young men's perceptions of masculinity and voluntary medical male circumcision. Findings from thematic analysis reveal that young men described several key characteristics of masculinity including responsibility, bravery and sexual attractiveness. For some young men, voluntary medical male circumcision has embedded itself into cultural norms of masculinity by being a step in the transition from boyhood to manhood and by being a marker of some of these masculine characteristics. In the case of voluntary medical male circumcision, there may be opportunities to integrate other programming that helps men transition into healthy adulthood.

The burden of unrecognised chronic kidney disease in patients with type 2 diabetes at a county hospital clinic in Kenya: implications to care and need for screening.

BACKGROUND: Chronic Kidney Disease (CKD) in patients with type 2 diabetes enhances the cardiovascular risk profiles and disease, and is a strong predictor of progression to end-stage kidney disease. Early diagnosis is encouraged for referral to specialist kidney care to initiate active management that would optimize outcomes including forestalling progression to end-stage kidney disease. This study was conducted in a regional referral public health facility in Central Kenya with a high prevalence of type 2 diabetes. It was aimed at finding out the burden of undiagnosed chronic kidney disease in their clinic of ambulatory patients with type 2 diabetes who dwell mainly in the rural area. METHODS: A cross-sectional study was conducted at the out-patient of Nyeri County hospital. A total of 385 patients were enrolled over 5 months. Informed consent was obtained and clinical evaluation was done, a spot sample of urine obtained for albuminuria and venous blood drawn for HbA1c, Lipids and serum creatinine. Estimated GFR (eGFR) was calculated using the Cockroft-Gault equation. Chronic kidney disease (CKD) was classified on KDIGO scale. Albuminuria was reported as either positive or negative. Descriptive statistics for data summary and regression analysis were employed on SPSS v23. RESULTS: A total of 385 participants were included in the study, 252 (65.5%) were females. There were 39.0% (95%CI 34.3-44.2) patients in CKD/KDIGO stages 3, 4 and 5 and 32.7% (95%CI, 27.8-37.4) had Albuminuria. The risk factors that were significantly associated with chronic kidney disease/KDIGO stages 3, 4 and 5 were: age > 50 years, long duration with diabetes > 5 years and hypertension. Employment and paradoxically, obesity reduced the odds of having CKD, probably as markers of better socio-economic status. CONCLUSION: Unrecognized CKD of KDIGO stages 3,4 and 5 occurred in over 30 % of the study patients. The risk factors of hypertension, age above 50, long duration of diabetes should help identify those at high risk of developing CKD, for screening and linkage to care. They are at high risk of progression to end-stage kidney disease and cardiovascular events. The imperative of screening for chronic kidney disease is availing care in publicly-funded hospitals.

Adequacy of control of cardiovascular risk factors in ambulatory patients with type 2 diabetes attending diabetes out-patients clinic at a county hospital, Kenya.

BACKGROUND: Type 2 diabetes is associated with substantial cardiovascular morbidity and mortality arising from the high prevalence of cardiovascular risk factors such as hypertension, dyslipidaemia, obesity, poor glycaemic control and albuminuria. Adequacy of control of these risk factors determines the frequency and outcome of cardiovascular events in the patients. Current clinical practice guidelines emphasize primary prevention of cardiovascular disease in type 2 diabetes. There is scarce data from the developing countries, Kenya included, on clinical care of patients with type 2 diabetes in the regions that are far away from tertiary health facilities. So we determined the adequacy of control of the modifiable risk factors: glycaemic control, hypertension, dyslipidemia, obesity and albuminuria in the study patients from rural and peri-urban dwelling. METHODS: This was a cross-sectional study on 385 randomly selected ambulatory patients with type 2 diabetes without overt complications. They were on follow up for at least 6 months at the Out-patient diabetes clinic of Nyeri County Hospital, a public health facility located in the central region of Kenya. RESULTS: Females were 65.5%. The study subjects had a mean duration of diabetes of 9.4 years, IQR of 3.0-14 years. Their mean age was 63.3 years, IQR of 56-71 years. Only 20.3% of our subjects had simultaneous optimal control of the three (3) main cardiovascular risk factors of hypertension, high LDL-C and hyperglycaemia at the time of the study. The prevalence of cardiovascular risk factors were as follows: HbA1c above 7% was 60.5% (95% CI, 55.6-65.5), hypertension, 49.6% of whom 76.6% (95% CI, 72.5-80.8) were poorly controlled. High LDL-Cholesterol above 2.0 mmol/L was found in 77.1% (95% CI 73.0-81.3) and Albuminuria occurred in 32.7% (95% CI 27.8-37.4). The prevalence of the other habits with cardiovascular disease risk were: excess alcohol intake at 26.5% (95% CI 27.8-37.4) and cigarette-smoking at 23.6%. A modest 23.4% of the treated patients with hypertension attained target blood pressure of <140/90 mmHg. Out of a paltry 12.5% of the statin-treated patients and others not actively treated, only 22.9% had LDL-Cholesterol of target <2.0 mmol/L. There were no obvious socio-demographic and clinical determinants of poor glycaemic control. However, old age above 50 yrs., longer duration with diabetes above 5 yrs. and advanced stages of CKD were significantly associated with hypertension. Female gender and age, statin non-use and socio-economic factor of employment were the significant determinants of high levels of serum LDL-cholesterol. CONCLUSION: The majority of the study patients attending this government-funded health facility had high prevalence of cardiovascular risk factors that were inadequately controlled. Therefore patients with type 2 diabetes should be risk-stratified by their age, duration of diabetes and cardiovascular risk factor loading. Consequently, composite risk factor reduction strategies are needed in management of these patients to achieve the desired targets safely. This would be achieved through innovative care systems and modes of delivery which would translate into maximum benefit of primary cardiovascular disease prevention in those at high risk. It is a desirable quality objective to have a higher proportion of the patients who access care benefiting maximally more than the numbers we are achieving now.

Mutation in IRF2BP2 is responsible for a familial form of common variable immunodeficiency disorder.

BACKGROUND: Genome-wide association studies have shown a pattern of rare copy number variations and single nucleotide polymorphisms in patients with common variable immunodeficiency disorder (CVID), which was recognizable by a support vector machine (SVM) algorithm. However, rare monogenic causes of CVID might lack such a genetic fingerprint. OBJECTIVE: We sought to identify a unique monogenic cause of familial immunodeficiency and evaluate the use of SVM to identify patients with possible monogenic disorders. METHODS: A family with multiple members with a diagnosis of CVID was screened by using whole-exome sequencing. The proband and other subjects with mutations associated with CVID-like phenotypes were screened through the SVM algorithm from our recent CVID genome-wide association study. RT-PCR, protein immunoblots, and in vitro plasmablast differentiation assays were performed on patient and control EBV lymphoblastoids cell lines. RESULTS: Exome sequencing identified a novel heterozygous mutation in IRF2BP2 (c.1652G>A:p.[S551N]) in affected family members. Transduction of the mutant gene into control human B cells decreased production of plasmablasts in vitro, and IRF2BP2 transcripts and protein expression were increased in proband versus control EBV-immortalized lymphoblastoid cell lines. The SVM algorithm categorized the proband and subjects with other immunodeficiency-associated gene variants in TACI, BAFFR, ICOS, CD21, LRBA, and CD27 as genetically dissimilar from polygenic CVID. CONCLUSION: A novel IRFBP2 mutation was identified in a family with autosomal dominant CVID. Transduction experiments suggest that the mutant protein has an effect on B-cell differentiation and is likely a monogenic cause of the family's CVID phenotype. Successful grouping by the SVM algorithm suggests that our family and other subjects with rare immunodeficiency disorders cluster separately and lack the genetic pattern present in polygenic CVID cases.

Mutations in PDGFRB cause autosomal-dominant infantile myofibromatosis.

Infantile myofibromatosis (IM) is a disorder of mesenchymal proliferation characterized by the development of nonmetastasizing tumors in the skin, muscle, bone, and viscera. Occurrence within families across multiple generations is suggestive of an autosomal-dominant (AD) inheritance pattern, but autosomal-recessive (AR) modes of inheritance have also been proposed. We performed whole-exome sequencing (WES) in members of nine unrelated families clinically diagnosed with AD IM to identify the genetic origin of the disorder. In eight of the families, we identified one of two disease-causing mutations, c.1978C>A (p.Pro660Thr) and c.1681C>T (p.Arg561Cys), in PDGFRB. Intriguingly, one family did not have either of these PDGFRB mutations but all affected individuals had a c.4556T>C (p.Leu1519Pro) mutation in NOTCH3. Our studies suggest that mutations in PDGFRB are a cause of IM and highlight NOTCH3 as a candidate gene. Further studies of the crosstalk between PDGFRB and NOTCH pathways may offer new opportunities to identify mutations in other genes that result in IM and is a necessary first step toward understanding the mechanisms of both tumor growth and regression and its targeted treatment.

Autism genome-wide copy number variation reveals ubiquitin and neuronal genes.

Autism spectrum disorders (ASDs) are childhood neurodevelopmental disorders with complex genetic origins. Previous studies focusing on candidate genes or genomic regions have identified several copy number variations (CNVs) that are associated with an increased risk of ASDs. Here we present the results from a whole-genome CNV study on a cohort of 859 ASD cases and 1,409 healthy children of European ancestry who were genotyped with approximately 550,000 single nucleotide polymorphism markers, in an attempt to comprehensively identify CNVs conferring susceptibility to ASDs. Positive findings were evaluated in an independent cohort of 1,336 ASD cases and 1,110 controls of European ancestry. Besides previously reported ASD candidate genes, such as NRXN1 (ref. 10) and CNTN4 (refs 11, 12), several new susceptibility genes encoding neuronal cell-adhesion molecules, including NLGN1 and ASTN2, were enriched with CNVs in ASD cases compared to controls (P = 9.5 x 10(-3)). Furthermore, CNVs within or surrounding genes involved in the ubiquitin pathways, including UBE3A, PARK2, RFWD2 and FBXO40, were affected by CNVs not observed in controls (P = 3.3 x 10(-3)). We also identified duplications 55 kilobases upstream of complementary DNA AK123120 (P = 3.6 x 10(-6)). Although these variants may be individually rare, they target genes involved in neuronal cell-adhesion or ubiquitin degradation, indicating that these two important gene networks expressed within the central nervous system may contribute to the genetic susceptibility of ASD.

Strong synaptic transmission impact by copy number variations in schizophrenia.

Schizophrenia is a psychiatric disorder with onset in late adolescence and unclear etiology characterized by both positive and negative symptoms, as well as cognitive deficits. To identify copy number variations (CNVs) that increase the risk of schizophrenia, we performed a whole-genome CNV analysis on a cohort of 977 schizophrenia cases and 2,000 healthy adults of European ancestry who were genotyped with 1.7 million probes. Positive findings were evaluated in an independent cohort of 758 schizophrenia cases and 1,485 controls. The Gene Ontology synaptic transmission family of genes was notably enriched for CNVs in the cases (P = 1.5 x 10(-7)). Among these, CACNA1B and DOC2A, both calcium-signaling genes responsible for neuronal excitation, were deleted in 16 cases and duplicated in 10 cases, respectively. In addition, RET and RIT2, both ras-related genes important for neural crest development, were significantly affected by CNVs. RET deletion was exclusive to seven cases, and RIT2 deletions were overrepresented common variant CNVs in the schizophrenia cases. Our results suggest that novel variations involving the processes of synaptic transmission contribute to the genetic susceptibility of schizophrenia.

Phylogeographic Assessment Reveals Geographic Sources of HIV-1 Dissemination Among Men Who Have Sex With Men in Kenya.

HIV-1 transmission dynamics involving men who have sex with men (MSM) in Africa are not well understood. We investigated the rates of HIV-1 transmission between MSM across three regions in Kenya: Coast, Nairobi, and Nyanza. We analyzed 372 HIV-1 partial pol sequences sampled during 2006-2019 from MSM in Coast (N = 178, 47.9%), Nairobi (N = 137, 36.8%), and Nyanza (N = 57, 15.3%) provinces in Kenya. Maximum-likelihood (ML) phylogenetics and Bayesian inference were used to determine HIV-1 clusters, evolutionary dynamics, and virus migration rates between geographic regions. HIV-1 sub-subtype A1 (72.0%) was most common followed by subtype D (11.0%), unique recombinant forms (8.9%), subtype C (5.9%), CRF 21A2D (0.8%), subtype G (0.8%), CRF 16A2D (0.3%), and subtype B (0.3%). Forty-six clusters (size range 2-20 sequences) were found-half (50.0%) of which had evidence of extensive HIV-1 mixing among different provinces. Data revealed an exponential increase in infections among MSM during the early-to-mid 2000s and stable or decreasing transmission dynamics in recent years (2017-2019). Phylogeographic inference showed significant (Bayes factor, BF > 3) HIV-1 dissemination from Coast to Nairobi and Nyanza provinces, and from Nairobi to Nyanza province. Strengthening HIV-1 prevention programs to MSM in geographic locations with higher HIV-1 prevalence among MSM (such as Coast and Nairobi) may reduce HIV-1 incidence among MSM in Kenya.

Quantifying rates of HIV-1 flow between risk groups and geographic locations in Kenya: A country-wide phylogenetic study.

In Kenya, HIV-1 key populations including men having sex with men (MSM), people who inject drugs (PWID) and female sex workers (FSW) are thought to significantly contribute to HIV-1 transmission in the wider, mostly heterosexual (HET) HIV-1 transmission network. However, clear data on HIV-1 transmission dynamics within and between these groups are limited. We aimed to empirically quantify rates of HIV-1 flow between key populations and the HET population, as well as between different geographic regions to determine HIV-1 'hotspots' and their contribution to HIV-1 transmission in Kenya. We used maximum-likelihood phylogenetic and Bayesian inference to analyse 4058 HIV-1 pol sequences (representing 0.3 per cent of the epidemic in Kenya) sampled 1986-2019 from individuals of different risk groups and regions in Kenya. We found 89 per cent within-risk group transmission and 11 per cent mixing between risk groups, cyclic HIV-1 exchange between adjoining geographic provinces and strong evidence of HIV-1 dissemination from (i) West-to-East (i.e. higher-to-lower HIV-1 prevalence regions), and (ii) heterosexual-to-key populations. Low HIV-1 prevalence regions and key populations are sinks rather than major sources of HIV-1 transmission in Kenya. Targeting key populations in Kenya needs to occur concurrently with strengthening interventions in the general epidemic.

Menstrual Needs and Associations with Sexual and Reproductive Risks in Rural Kenyan Females: A Cross-Sectional Behavioral Survey Linked with HIV Prevalence.

BACKGROUND: Females in low and middle income countries (LMICs) have difficulty coping with menstrual needs, but few studies have examined the social or health implications of these needs. METHODS: Responses from 3418 menstruating females aged 13-29 years were extracted from an HIV and behavioral risks cross-sectional survey conducted in rural western Kenya. We examined sanitary products used, provision of products from sexual partners or from transactional sex, and demographic and sexual exposures. RESULTS: Overall, 75% of females reported using commercial pads and 25% used traditional materials such as cloth or items like paper or tissue, with 10% of girls <15 years old depending on makeshift items. Two-thirds of females with no education relied on traditional items. Having attended secondary school increased the odds of using commercial pads among married (adjusted odds ratios [AOR] 4.8, 95% confidence interval [CI] 3.25-7.12) and single females (AOR 2.17, 95% CI 1.04-4.55). Married females had lower odds of pad use if they reported early (<12 years of age) compared with later (≥18 years) sexual debut (64% vs. 78%, AOR 0.45, 95% CI 0.21-0.97). Two-thirds of pad users received them from sexual partners. Receipt was lower among married females if partners were violent (AOR 0.67, 95% CI 0.53-0.85). Receipt among single females was higher if they had two or more sexual partners in the past year (AOR 2.11, 95% CI 1.04-4.29). Prevalence of engaging in sex for money to buy pads was low (1.3%); however, 10% of 15-year-olds reported this, with girls ≤15 having significantly higher odds compared with females over 15 (AOR 2.84, 95% CI 0.89-9.11). The odds of having transactional sex for pads was higher among females having two or more partners in the past 12 months (AOR 4.86, 95% CI 2.06-11.43). CONCLUSIONS: Menstrual needs of impoverished females in rural LMICs settings likely leads to increased physical and sexual harms. Studies are required to strengthen knowledge and to evaluate interventions to reduce these harms.

AGC1 Deficiency Causes Infantile Epilepsy, Abnormal Myelination, and Reduced N-Acetylaspartate.

BACKGROUND: Whole exome sequencing (WES) offers a powerful diagnostic tool to rapidly and efficiently sequence all coding genes in individuals presenting for consideration of phenotypically and genetically heterogeneous disorders such as suspected mitochondrial disease. Here, we report results of WES and functional validation in a consanguineous Indian kindred where two siblings presented with profound developmental delay, congenital hypotonia, refractory epilepsy, abnormal myelination, fluctuating basal ganglia changes, cerebral atrophy, and reduced N-acetylaspartate (NAA). METHODS: Whole blood DNA from one affected and one unaffected sibling was captured by Agilent SureSelect Human All Exon kit and sequenced on the Illumina HiSeq2000. Mutations were validated by Sanger sequencing in all family members. Protein from wild-type and mutant fibroblasts was isolated to assess mutation effects on protein expression and enzyme activity. RESULTS: A novel SLC25A12 homozygous missense mutation, c.1058G>A; p.Arg353Gln, segregated with disease in this kindred. SLC25A12 encodes the neuronal aspartate-glutamate carrier 1 (AGC1) protein, an essential component of the neuronal malate/aspartate shuttle that transfers NADH and H(+) reducing equivalents from the cytosol to mitochondria. AGC1 activity enables neuronal export of aspartate, the glial substrate necessary for proper neuronal myelination. Recombinant mutant p.Arg353Gln AGC1 activity was reduced to 15% of wild type. One prior reported SLC25A12 mutation caused complete loss of AGC1 activity in a child with epilepsy, hypotonia, hypomyelination, and reduced brain NAA. CONCLUSIONS: These data strongly suggest that SLC25A12 disease impairs neuronal AGC1 activity. SLC25A12 sequencing should be considered in children with infantile epilepsy, congenital hypotonia, global delay, abnormal myelination, and reduced brain NAA.

Identification of rare DNA sequence variants in high-risk autism families and their prevalence in a large case/control population.

BACKGROUND: Genetics clearly plays a major role in the etiology of autism spectrum disorders (ASDs), but studies to date are only beginning to characterize the causal genetic variants responsible. Until recently, studies using multiple extended multi-generation families to identify ASD risk genes had not been undertaken. METHODS: We identified haplotypes shared among individuals with ASDs in large multiplex families, followed by targeted DNA capture and sequencing to identify potential causal variants. We also assayed the prevalence of the identified variants in a large ASD case/control population. RESULTS: We identified 584 non-conservative missense, nonsense, frameshift and splice site variants that might predispose to autism in our high-risk families. Eleven of these variants were observed to have odds ratios greater than 1.5 in a set of 1,541 unrelated children with autism and 5,785 controls. Three variants, in the RAB11FIP5, ABP1, and JMJD7-PLA2G4B genes, each were observed in a single case and not in any controls. These variants also were not seen in public sequence databases, suggesting that they may be rare causal ASD variants. Twenty-eight additional rare variants were observed only in high-risk ASD families. Collectively, these 39 variants identify 36 genes as ASD risk genes. Segregation of sequence variants and of copy number variants previously detected in these families reveals a complex pattern, with only a RAB11FIP5 variant segregating to all affected individuals in one two-generation pedigree. Some affected individuals were found to have multiple potential risk alleles, including sequence variants and copy number variants (CNVs), suggesting that the high incidence of autism in these families could be best explained by variants at multiple loci. CONCLUSIONS: Our study is the first to use haplotype sharing to identify familial ASD risk loci. In total, we identified 39 variants in 36 genes that may confer a genetic risk of developing autism. The observation of 11 of these variants in unrelated ASD cases further supports their role as ASD risk variants.

Whole-genome sequencing in an autism multiplex family.

BACKGROUND: Autism spectrum disorders (ASDs) represent a group of childhood neurodevelopmental disorders that affect 1 in 88 children in the US. Previous exome sequencing studies on family trios have implicated a role for rare, de-novo mutations in the pathogenesis of autism. METHODS: To examine the utility of whole-genome sequencing to identify inherited disease candidate variants and genes, we sequenced two probands from a large pedigree, including two parents and eight children. We evaluated multiple analytical strategies to identify a prioritized list of candidate genes. RESULTS: By assuming a recessive model of inheritance, we identified seven candidate genes shared by the two probands. We also evaluated a different analytical strategy that does not require the assumption of disease model, and identified a list of 59 candidate variants that may increase susceptibility to autism. Manual examination of this list identified ANK3 as the most likely candidate gene. Finally, we identified 33 prioritized non-coding variants such as those near SMG6 and COQ5, based on evolutionary constraint and experimental evidence from ENCODE. Although we were unable to confirm rigorously whether any of these genes indeed contribute to the disease, our analysis provides a prioritized shortlist for further validation studies. CONCLUSIONS: Our study represents one of the first whole-genome sequencing studies in autism leveraging a large family-based pedigree. These results provide for a discussion on the relative merits of finding de-novo mutations in sporadic cases versus finding inherited mutations in large pedigrees, in the context of neuropsychiatric and neurodevelopmental diseases.

Whole-genome DNA/RNA sequencing identifies truncating mutations in RBCK1 in a novel Mendelian disease with neuromuscular and cardiac involvement.

BACKGROUND: Whole-exome sequencing has identified the causes of several Mendelian diseases by analyzing multiple unrelated cases, but it is more challenging to resolve the cause of extremely rare and suspected Mendelian diseases from individual families. We identified a family quartet with two children, both affected with a previously unreported disease, characterized by progressive muscular weakness and cardiomyopathy, with normal intelligence. During the course of the study, we identified one additional unrelated patient with a comparable phenotype. METHODS: We performed whole-genome sequencing (Complete Genomics platform), whole-exome sequencing (Agilent SureSelect exon capture and Illumina Genome Analyzer II platform), SNP genotyping (Illumina HumanHap550 SNP array) and Sanger sequencing on blood samples, as well as RNA-Seq (Illumina HiSeq platform) on transformed lymphoblastoid cell lines. RESULTS: From whole-genome sequence data, we identified RBCK1, a gene encoding an E3 ubiquitin-protein ligase, as the most likely candidate gene, with two protein-truncating mutations in probands in the first family. However, exome data failed to nominate RBCK1 as a candidate gene, due to poor regional coverage. Sanger sequencing identified a private homozygous splice variant in RBCK1 in the proband in the second family, yet SNP genotyping revealed a 1.2Mb copy-neutral region of homozygosity covering RBCK1. RNA-Seq confirmed aberrant splicing of RBCK1 transcripts, resulting in truncated protein products. CONCLUSIONS: While the exact mechanism by which these mutations cause disease is unknown, our study represents an example of how the combined use of whole-genome DNA and RNA sequencing can identify a disease-predisposing gene for a novel and extremely rare Mendelian disease.

Baseline findings of an HIV incidence cohort study to prepare for future HIV prevention clinical trials in Kisumu, Kenya.

INTRODUCTION: In an analysis of baseline findings of an HIV incidence cohort study, an assessment was made of HIV prevalence among persons presenting for enrollment and any differences in demographic characteristics between persons not enrolled compared to those enrolled. We also described and compared HIV risk behaviors in males and females enrolled in the study. METHODOLOGY: A computer-assisted survey was administered to collect baseline demographic and HIV risk data from 1,277 men and women aged 18-34 years. Testing for HIV and other sexually transmitted infections (STI) was conducted. Out of 1,277 persons prescreened for eligibility, 625 were enrolled. RESULTS: HIV prevalence of all persons who completed screening was 14.8% (females: 21.1%; males: 8.1%). The odds of being enrolled in the study were higher for persons 18-24 years compared to those 30-34 years of age [adjusted odds ratio (AOR)=2.18, CI=1.13, 4.21] and males compared to females [AOR=2.07, CI=1.43, 2.99]. Among those enrolled in the study, the most prevalent HIV risk behaviors were unprotected sex (49%), alcohol use (45%), and transactional sex (30%) in the last three months. Compared to females, a significantly greater proportion of males reported using any alcohol or recreational drug in the last three months, a history of oral sex, sex with partner other than a spouse or main partner, ever having a blood transfusion, ever being treated for an STI, and having knowledge of their last HIV test result. CONCLUSION: The Kisumu Field Station successfully recruited individuals with HIV risk characteristics for the HIV incidence cohort study.