An Eco-Benign Biomimetic Approach for the Synthesis of Ni/ZnO Nanocomposite: Photocatalytic and Antioxidant Activities.

With the increasing demand for wastewater treatment and multidrug resistance among pathogens, it was necessary to develop an efficient catalyst with enhanced photocatalytic and antibacterial applications. The present study proposes a facile and green strategy for synthesizing zinc oxide (ZnO) decorated nickel (Ni) nanomaterials. The synthesized Ni/ZnO nanocomposite displays a high crystallinity and spherical morphology, which was systematically characterized by XRD, SEM, FT-IR, UV-visible spectroscopy, EDX, HRTEM, and XPS techniques. In addition, the bacteriological tests indicated that Ni/ZnO nanocomposite exhibits potent antibacterial activity against human pathogens, i.e., Pseudomonas aeruginosa (P. aeruginosa), Staphylococcus aureus (S. aureus), and Escherichia coli (E. coli). The inhibition zone observed in light and dark conditions for E. coli was 16 (±0.3) mm and 8 (±0.4) mm, respectively, which confirms the high efficacy of the nanocomposite in the presence of light compared to dark conditions. The detailed inhibition mechanism of said bacterium and damage were also studied through fluorescence spectroscopy and SEM analysis, respectively. Evaluation of antioxidant activity based on free radical scavenging activity revealed that the Ni/ZnO nanocomposite effectively scavenges DPPH. In the photocatalytic performance, the Ni/ZnO nanocomposite exhibited a remarkable degradation ability under the optimized condition, which was attributed to their controllable size, high surface area, and exceptional morphology. Good selectivity, high photodegradation, and antibacterial activities and satisfactory hemolytic behavior of the as-prepared nanocomposite make them able to become a potential candidate for superior biological performance and environmental remediation.

Risk of heavy metals accumulation in soil and wheat grains with waste water irrigation under different NPK levels in alkaline calcareous soil.

A field study was conducted on the reuse of wastewater from Mardan city to evaluate its risk of contaminating soil and wheat grains at different NPK levels. Three irrigation sources i.e. waste water (WW), canal water (CW) and alternate waste + canal water (WW+CW) were applied to wheat (cv Atta Habib 2010) grown at 0, 50, 75 and 100% NPK levels of 120:90:60 kg N:P2O5:K2O ha-1 at Palatoo Research Farm, Amir Muhammad Khan Campus, Mardan during 2015.The results showed higher grain and biomass yields in WW irrigated plots as compared to CW at NPK levels up to 50% of recommending dose revealing supplementing nutrient requirements in deficient conditions. However, irrigation of WW at higher NPK levels especially at or beyond 75% of recommended dose tended to reduce the crop yield that could be associated with heavy metals toxicity and nutritional imbalances. The use of WW substantially increased AB-DTPA extractable Zn, Mn, Pb, Ni and Cd indicating a potential threat to soil contamination. Similarly, WW irrigated wheat had higher concentrations of these heavy metals as compared to CW which limits its use for production purposes without any remediation measures. The alternate use of CW and WW as revealed by its comparative lower contamination in soil and wheat than sole WW could be one of the possible solutions and may increase the time required for threshold soil contamination.

Heavy metals immobilization and improvement in maize (Zea mays L.) growth amended with biochar and compost.

Soil with heavy metals contamination, mainly lead (Pb), cadmium (Cd), and chromium (Cr) is a progressively worldwide alarming environmental problem. Recently, biochar has been used as a soil amendment to remediate contaminated soils, but little work has been done to compare with other organic amendments like compost. We investigated biochar and compost's comparative effect on Pb, Cd, and Cr immobilization in soil, photosynthesis, and growth of maize plants. Ten kg soil was placed in pots and were spiked with Pb, Cd, and Cr at concentrations 20, 10, 20 mg kg-1. The biochar and compost treatments included 0, 0.5, 1, 2, and 4% were separately applied to the soil. The crop from pots was harvested after 60 days. The results show that the highest reduction of AB-DTPA extractable Pb, Cd, and Cr in soil was 79%, 61% and 78% with 4% biochar, followed by 61%, 43% and 60% with 4% compost compared to the control, respectively. Similarly, the highest reduction in shoot Pb, Cd, and Cr concentration was 71%, 63% and 78%with 4% biochar, followed by 50%, 50% and 71% with 4% compost than the control, respectively. The maximum increase in shoot and dry root weight, total chlorophyll contents, and gas exchange characteristics were recorded with 4% biochar, followed by 4% compost than the control. The maximum increase in soil organic matter and total nitrogen (N) was recorded at 4% biochar application while available phosphorus and potassium in the soil at 4% compost application. It is concluded that both biochar and compost decreased heavy metals availability in the soil, reducing toxicity in the plant. However, biochar was most effective in reducing heavy metals content in soil and plant compared to compost. In the future, more low-cost, eco-friendly soil remediation methods should be developed for better soil health and plant productivity.

Investigating the degradation behavior of Cypermethrin (CYP) and Chlorpyrifos (CPP) in peach orchard soils using organic/inorganic amendments.

Reducing toxic effects of pesticide residues in agricultural soils through organic amendments is an eco-friendly technique. Cypermethrin (CYP) and Chlorpyrifos (CPP) are widely used pesticides in peach growing orchards in Swat valley of Pakistan. The aim of the current study was to investigate the degradation behavior of CYP and CPP in soil by the application of different combination of organic/inorganic amendments. A total of 36 soil samples were used in the current incubation study which was collected from 4 peach orchards in district Swat, Khyber Pakhtunkhwa (KPK), Pakistan. Different amendments including urea, farm yard manure (FYM) and saprofil were applied alone and in various combinations. The initial concentrations of CYP and CPP in the tested soil was range from 0.94 to 4.8 mg kg-1 and 0.024 to 4.12 to mg kg-1. Soil samples were taken at 5, 15, 30 and 45 days after exposure to different treatments. The extraction of pesticides from soils was done through quick, easy, cheap, effective, rugged, and safe (QuEChERS) extraction method. Soils amended with urea, FYM and saprofil individually and in combinations significantly reduced the concentrations of CYP and CPP. However, the concentration of CYP (24.6) and CPP (27.0) in soil showed higher reduction through the application of FYM. While the concentrations of CYP and CPP were declined with the 5, 15, 30 and 45 days intervals, however, reduction at day 30 and 45 was faster for CYP (16.7 to 8.46) than CPP (20.2 to 12.3). At day 5 and 15, the CYP (42.5 to 30.7) was slightly lower than CPP (42.9 to 32.7).The highest half-life value (t ½) of CYP was in control treatment (32 days) and the shortest was soil amended with FYM (18.6 days). While the longest half-life value (t ½) of CPP was maximum in control treatment (42 days) and the minimum was in FYM (22 days). Based on our findings, it was concluded that soil application of FYM is recommended for the degradation of CYP and CPP.

Comparing the phosphorus use efficiency of pre-treated (organically) rock phosphate with soluble P fertilizers in maize under calcareous soils.

In calcareous soils, phosphorus (P) availability to plant is impaired due to the formation of insoluble complexes with calcium and magnesium. Therefore, this study was executed to compare the P use efficiency (PUE) of four different P sources [rock phosphate (RP), acidulated rock phosphate (ARP), single super phosphate (SSP) and di ammonium phosphate (DAP)] alone or pre-treated with organic amendments (farm yard manure (FYM) enriched compost, simple compost and humic acid (HA)) along with control in maize crop under calcareous soils. All treatments irrespective of P sources received 90 kg P2O5 ha-1. Phosphorus application regardless of its sources and combination with organic amendments significantly improved maize growth, yield as well as P uptake and PUE. Rock phosphate when applied alone was recorded inferior but its performance significantly improved with compost or its pre-addition with FYM and HA, that further enhanced upon acidulation. Maize grain yield increased by 21, 22.2, 67.9 and 94% with RP, ARP, ARP enriched compost and ARP+ compost respectively, over control. Similarly, PUE of DAP improved from 31.7 to 43.1 and 39 with sample and enriched compost correspondingly. Post-harvest soil and grain P were at par for SSP, ARP and DAP alone or in conjugation with organic amendments when averaged across the amendments. These results suggested that pretreatment of P sources with organic amendments is an economical and more feasible approach to improve maize yield and PUE. Moreover, on-farm acidulation of RP may give at par results with SSP and DAP with cheaper rate and hence recommended for P management in maize in alkaline calcareous soils.

Phosphorus Nutrient Management through Synchronization of Application Methods and Rates in Wheat and Maize Crops.

Management of inorganic fertilizer is very important to obtain maximum crop yield and improved nutrient use efficiency in cereal crops. Fixation of phosphatic fertilizers in alkaline soils due to calcareousness is one of the major hurdles. It induces phosphorus nutritional stress that can decrease the yield of maize and wheat. Selection of a suitable application method and proper stage of crop for phosphorus (P) fertilizer has prime importance in better uptake of P and crop production. Among different application methods, soil and foliar application are widely adopted. In wheat and maize, knee height + tasseling and stem elongation + booting are critical stages towards P deficiency. That is why field trials were conducted to evaluate the supplemental effect of foliar P on maize and wheat yields. For that, 144 mM KH2PO4 was applied as foliar at knee height + tasseling and stem elongation + boot stages in maize and wheat, respectively. Soil application of 0, 20, 40 and 60 kg P ha-1 was done through broadcast and band methods. Results showed that foliar spray of 144 mM KH2PO4 at knee height + tasseling and stem elongation + boot stages in wheat and maize significantly enhanced grains yield and phosphorus use efficiency (PUE) where P was applied as banding or broadcast at the time of sowing. A significant decreasing trend in response to increasing soil P levels validated the efficacious role and suitability of foliar P. In conclusion, the use of P as foliar at knee height + tasseling and stem elongation + boot stages is an efficacious way to manage P fertilizer.

Consanguinity, the driving force behind inheritance of HbS-ß thalassemia in Southern Districts of KP.

OBJECTIVE: To determine the clinical, haematological and genetic factors responsible for variable phenotypes of sickle haemoglobin, sickle haemoglobin-beta, and beta-thalassemia patients. METHODS: The study was conducted in Bannu, Lakki, Tank and Dera Ismail Khan districts of Khyber Pakhtunkhwa province of Pakistan from September 2016 to November 2017, and comprised sickle haemoglobin, sickle haemoglobin-beta, and beta-thalassemia patients. Clinical, haematological and genetic determinants were evaluated using haemoglobin electrophoresis and allele-specific primers through polymerase chain reaction to determine alpha and beta thalassemia, and CgT substitution at position -158 (referred to as Xmn-I polymorphism) in gamma-globin gene. Data was analysed using SPSS 20. RESULTS: Eight b-thalassemia mutations were identified that included IVS I-5(G C), codon 8/9 (+G), codon 30 (G C), -88 (C T), Cap+1(A G), codon 41/42 (-TCTT), IVS I-1(G T) and codon 16(-C). Codon 30 (G C) and -88 (C T) were found only in Pashtoon subjects, Cap+1(A G) and IVS I-1(G T) in Balochi subjects, while 75% of IVS I-5(G C) mutation cases were found in Punjabi ethnic group. In the Pashtoon group, 13 sickle haemoglobin homozygous patients were identified for the first time. Both alpha thalassemia and Xmn-I polymorphism in homozygous condition were common among those with mild phenotype. CONCLUSION: Phenotypic expression of sickle haemoglobin beta thalassemia was found to be extremely variable and alpha thalassemia and Xmn-I polymorphism in homozygous condition were found to be additional genetic modifiers of the disease.

Prevalence of Epstein­Barr Virus Genotypes in Pakistani Lymphoma Patients

The Epstein-Barr virus (EBV) is a herpesvirus infecting more than 90% of the human population. The tropism of EBV for B lymphocytes is evidenced in its association with many lymphoproliferative disorders. Different types of EBV (EBV-1 and EBV-2), classified on the basis of EBV nuclear antigen-2 (EBNA-2) genotyping, have been reported in benign and malignant pathologies, but there is almost no information about their frequency in the Pakistani population. The aim of this study was to determine the frequency and distribution of EBNA-2-based EBV genotypes in lymphoma patients. Genomic DNA was extracted from formalin-fixed paraffin embedded (FFPE) tissue samples obtained from 73 EBV-DNA-positive lymphoma patients. The ß-globin gene was amplified to assess the presence and quality of cellular DNA from all samples. EBER-1 DNA was detected by PCR to confirm EBV presence in tissue samples. EBNA-1 mRNA relative quantification done by quantitative PCR substantiated EBNA-1 mRNA overexpression in 43.8% of EBV-positive cases in comparison to EBV-positive control cell line. EBNA-2 genotyping was done by nested PCR. Among typable samples, EBV-1 was found in 90.7% of samples while EBV-2 was present in 9.3% cases. These results show that EBV-1 was the most prevalent type in the lymphoma population of Pakistan. This epidemiology of EBV in Pakistani lymphoma patients represents an important first step in using EBV for prognosis and monitoring treatment response.

A disease-causing novel missense mutation in the ST14 gene underlies autosomal recessive ichthyosis with hypotrichosis syndrome in a consanguineous family.

Autosomal recessive ichthyosis with hypotrichosis (ARIH; MIM 602400) syndrome is characterized by diffused congenital ichthyosis and generalized non-scarring hypotrichosis. The underlying genetic cause of ARIH syndrome has been associated with sequence variants of the gene ST14, encoding type II transmembrane serine protease matriptase, which maps to chromosome 11q24.3. The current report aimed to investigate the clinical features and genetic cause of ARIH syndrome in a large consanguineous family of Pakistani origin. The technique of homozygosity mapping with highly polymorphic microsatellite markers was employed to establish linkage within the family. Sanger sequencing of exons and intron-exon boundaries of ST14 was performed to identify the potential pathogenic sequence variants, followed by structural analysis of the mutated protein. Linkage was established to chromosome 11q24.3, comprising the gene ST14. Sequence analysis led to the identification of a novel homozygous missense variant (c.1315G>A, p.Gly439Ser) in the ST14 gene that co-segregated with the disease phenotype in all affected members. Homology modelling and molecular docking analysis of ST14 with wild-type TMEFF1 protein was performed which revealed that glycine at position 439 is crucial for maintaining normal structural confirmation and interaction with the EGF domain of TMEFF1 protein. Taken together, the data strongly advocate this ST14 variant as the underlying genetic cause of ARIH syndrome in this first reported affected family from Pakistan. Moreover, the present study adds to the spectrum of mutations in the ST14 gene, implicating them in the pathogenesis of ARIH syndrome.

A novel homozygous variant in BMPR1B underlies acromesomelic dysplasia Hunter-Thompson type.

Acromesomelic dysplasia is genetically heterogeneous group of skeletal disorders characterized by short stature and acromelia and mesomelia of limbs. Acromesomelic dysplasia segregates in an autosomal recessive pattern and is caused by biallelic sequence variants in three genes (NPR2, GDF5, and BMPR1B). A consanguineous family of Pakistani origin segregating a subtype of acromesomelic dysplasia called Hunter-Thompson was clinically and genetically evaluated. Genotyping of microsatellite markers and linkage analysis revealed a 7.78 Mb homozygous region on chromosome 4q22.3, which harbors BMPR1B. Sequence analysis of the gene revealed a novel homozygous missense variant (c.1190T > G, p.Met397Arg) that segregates with the disease phenotype within the family and produced a Logarithm of odds (LOD) score of 3.9 with the disease phenotype. This study reports on the first familial case of acromesomelic dysplasia Hunter-Thompson type. It is also the first report of BMPR1B underlying the etiology of acromesomelic dysplasia Hunter-Thompson type.

Role of Streptomyces pactum in phytoremediation of trace elements by Brassica juncea in mine polluted soils.

The industrial expansion, smelting, mining and agricultural practices have increased the release of toxic trace elements (TEs) in the environment and threaten living organisms. The microbe-assisted phytoremediation is environmentally safe and provide an effective approach to remediate TEs contaminated soils. A pot experiment was conducted to test the potential of an Actinomycete, subspecies Streptomyces pactum (Act12) along with medical stone compost (MSC) by growing Brassica juncea in smelter and mines polluted soils of Feng County (FC) and Tongguan (TG, China), respectively. Results showed that Zn (7, 28%), Pb (54, 21%), Cd (16, 17%) and Cu (8, 10%) uptake in shoot and root of Brassica juncea was pronounced in FC soil. Meanwhile, the Zn (40, 14%) and Pb (82, 15%) uptake in the shoot and root were also increased in TG soil. Shoot Cd uptake remained below detection, while Cu decreased by 52% in TG soil. The Cd and Cu root uptake were increased by 17% and 33%, respectively. Results showed that TEs uptake in shoot increased with increasing Act12 dose. Shoot/root dry biomass, chlorophyll and carotenoid content in Brassica juncea were significantly influenced by the application of Act12 in FC and TG soil. The antioxidant enzymatic activities (POD, PAL, PPO and CAT) in Brassica juncea implicated enhancement in the plant defense mechanism against the TEs induced stress in contaminated soils. The extraction potential of Brasssica was further evaluated by TF (translocation factor) and MEA (metal extraction amount). Based on our findings, further investigation of Act12 assisted phytoremediation of TEs in the smelter and mines polluted soil and hyperaccumulator species are suggested for future studies.

A novel homozygous variant in the SMOC1 gene underlying Waardenburg anophthalmia syndrome.

BACKGROUND: Waardenburg anophthalmia syndrome (WAS), also known as ophthalmo-acromelic syndrome or anophthalmia-syndactyly, is a rare congenital disorder that segregates in an autosomal recessive pattern. Clinical features of the syndrome include malformation of the eyes and the skeleton. Mostly, WAS is caused by mutations in the SMOC-1 gene. MATERIALS AND METHODS: The present report describes a large consanguineous family of Pakistani origin segregating Waardenburg anophthalmia syndrome in an autosomal recessive pattern. Genotyping followed by Sanger sequencing was performed to search for a candidate gene. RESULTS: SNP genotyping using AffymetrixGeneChip Human Mapping 250K Nsp array established a single homozygous region among affected members on chromosome 14q23.1-q24.3 harboring the SMOC1 gene. Sequencing of the gene revealed a novel homozygous missense mutation (c.812G>A; p.Cys271Tyr) in the family. CONCLUSION: This is the first report of Waardenburg anophthalmia syndrome caused by a SMOC1 variant in a Pakistani population. The mutation identified in the present investigation extends the body of evidence implicating the gene SMOC-1 in causing WAS.

Synthesis of manganese incorporated hierarchical mesoporous silica nanosphere with fibrous morphology by facile one-pot approach for efficient catalytic ozonation.

Manganese incorporated fibrous silica nanosphere (MnOx-0.013/KCC-1) was synthesized by one step hydrothermal method for the first time and its catalytic activity for ozonation of oxalic acid was studied. For comparison, manganese loaded MCM-41 (MnOx-0.013/MCM-41) was prepared by impregnation method. Various characterizations showed that the morphological, structural and textural properties of MnOx-0.013/KCC-1 were well preserved. Ozonation and catalytic ozonation by MnOx-0.013/KCC-1 and MnOx-0.013/MCM-41 led to 4, 85 and 60% reduction in TOC respectively. Furthermore, 0.05 and 1.2mgL(-1) leaching of Mn was detected from MnOx-0.013/KCC-1 and MnOx-0.013/MCM-41, which are approximately 2.0 and 42.0% of the total Mn present in MnOx-0.013/KCC-1 and MnOx-0.013/MCM-41 respectively. The high catalytic activity was attributed to the generation of hydroxyl radical. Surface hydroxyl groups investigated by using phosphates and ATR-FTIR were believed to be the active sites. Our proposed method of synthesis can be generalized for the synthesis of other metal oxides incorporated fibrous silica for environmental catalysis and other catalytic reactions.

Autosomal-Recessive Hearing Impairment Due to Rare Missense Variants within S1PR2.

The sphingosine-1-phosphate receptors (S1PRs) are a well-studied class of transmembrane G protein-coupled sphingolipid receptors that mediate multiple cellular processes. However, S1PRs have not been previously reported to be involved in the genetic etiology of human traits. S1PR2 lies within the autosomal-recessive nonsyndromic hearing impairment (ARNSHI) locus DFNB68 on 19p13.2. From exome sequence data we identified two pathogenic S1PR2 variants, c.323G>C (p.Arg108Pro) and c.419A>G (p.Tyr140Cys). Each of these variants co-segregates with congenital profound hearing impairment in consanguineous Pakistani families with maximum LOD scores of 6.4 for family DEM4154 and 3.3 for family PKDF1400. Neither S1PR2 missense variant was reported among ∼120,000 chromosomes in the Exome Aggregation Consortium database, in 76 unrelated Pakistani exomes, or in 720 Pakistani control chromosomes. Both DNA variants affect highly conserved residues of S1PR2 and are predicted to be damaging by multiple bioinformatics tools. Molecular modeling predicts that these variants affect binding of sphingosine-1-phosphate (p.Arg108Pro) and G protein docking (p.Tyr140Cys). In the previously reported S1pr2(-/-) mice, stria vascularis abnormalities, organ of Corti degeneration, and profound hearing loss were observed. Additionally, hair cell defects were seen in both knockout mice and morphant zebrafish. Family PKDF1400 presents with ARNSHI, which is consistent with the lack of gross malformations in S1pr2(-/-) mice, whereas family DEM4154 has lower limb malformations in addition to hearing loss. Our findings suggest the possibility of developing therapies against hair cell damage (e.g., from ototoxic drugs) through targeted stimulation of S1PR2.

Novel mutations in the genes TGM1 and ALOXE3 underlying autosomal recessive congenital ichthyosis.

BACKGROUND: Ichthyoses are clinically characterized by scaling or hyperkeratosis of the skin or both. It can be an isolated condition limited to the skin or appear secondarily with involvement of other cutaneous or systemic abnormalities. METHODS: The present study investigated clinical and molecular characterization of three consanguineous families (A, B, C) segregating two different forms of autosomal recessive congenital ichthyosis (ARCI). Linkage in three consanguineous families (A, B, C) segregating two different forms of ARCI was searched by typing microsatellite and single nucleotide polymorphism marker analysis. Sequencing of the two genes TGM1 and ALOXE3 was performed by the dideoxy chain termination method. RESULTS: Genome-wide linkage analysis established linkage in family A to TGM1 gene on chromosome 14q11 and in families B and C to ALOXE3 gene on chromosome 17p13. Subsequently, sequencing of these genes using samples from affected family members led to the identification of three novel mutations: a missense variant p.Trp455Arg in TGM1 (family A); a nonsense variant p.Arg140* in ALOXE3 (family B); and a complex rearrangement in ALOXE3 (family C). CONCLUSION: The present study further extends the spectrum of mutations in the two genes involved in causing ARCI. Characterizing the clinical spectrum resulting from mutations in the TGM1 and ALOXE3 genes will improve diagnosis and may direct clinical care of the family members.

Mutations in the lipase-H gene causing autosomal recessive hypotrichosis and woolly hair.

Hypotrichosis is characterised by sparse scalp hair, sparse to absent eyebrows and eyelashes, or absence of hair from other parts of the body. In few cases, the condition is associated with tightly curled woolly scalp hair. The present study searched for disease-causing sequence variants in the genes in four Pakistani lineal consanguineous families exhibiting features of hypotrichosis or woolly hair. A haplotype analysis established links in all four families to the LIPH gene located on chromosome 3q27.2. Subsequently, sequencing LIPH identified a novel non-sense mutation (c.328C>T; p.Arg110*) in one and a previously reported 2-bp deletion mutation (c.659_660delTA, p.Ile220ArgfsX29) in three other families.

In silico analysis of missense mutations in LPAR6 reveals abnormal phospholipid signaling pathway leading to hypotrichosis.

Autosomal recessive hypotrichosis is a rare genetic irreversible hair loss disorder characterized by sparse scalp hair, sparse to absent eyebrows and eyelashes, and sparse axillary and body hair. The study, presented here, established genetic linkage in four families showing similar phenotypes to lysophosphatidic acid receptor 6 (LPAR6) gene on chromosome 13q14.11-q21.32. Subsequently, sequence analysis of the gene revealed two previously reported missense mutations including p.D63V in affected members of one and p.I188F in three other families. Molecular modeling and docking analysis was performed to investigate binding of a ligand oleoyl-L-alpha-lysophosphatidic acid (LPA) to modeled protein structures of normal and mutated (D63V, G146R, I188F, N248Y, S3T, L277P) LPAR6 receptors. The mutant receptors showed a complete shift in orientation of LPA at the binding site. In addition, hydropathy analysis revealed a significant change in the membrane spanning topology of LPAR6 helical segments. The present study further substantiated involvement of LPAR6-LPA signaling in the pathogenesis of hypotrichosis/woolly hair and provided additional insight into the molecular mechanism of hair development.

A novel missense mutation in MSX1 underlies autosomal recessive oligodontia with associated dental anomalies in Pakistani families.

Tooth agenesis constitutes the most common anomaly of dental development in humans. In the majority of familial cases of hypodontia alone or in association with other anomalies, the mode of inheritance is autosomal dominant. In the present study, we have identified two distantly related consanguineous Pakistani kindreds with an autosomal recessive form of oligodontia with associated dental anomalies. Locus in this case has been mapped on chromosome 4p16.1-p16.3. The maximum two-point LOD score of 2.85 (theta=0.0) was obtained at markers D4S2925 and D4S2285. A maximum multipoint LOD score exceeding 4 was obtained at the same markers. Recombination events observed in affected individuals localized the disease locus between markers D4S412 and D4S2935, spanning a 9.24-cM region on chromosome 4p16.1-p16.3. Sequence analysis of candidate gene MSX1 revealed a novel recessive missense mutation resulting in substitution of alanine to threonine amino acid (p. A219T), located in the MSX1 homeodomain, which is important for DNA binding and protein-protein interaction. The mutation, p. A219T, is the first recessive mutation identified in MSX1.