Antlions, a group of popular predatory insects, comes under the family Myrmeleontidae and the order Neuroptera. Both adults and larvae are predatory in nature. The proper identification of an organism is a key factor in understanding its biology. Due to their unique biology and mysterious living antlions generally remain under-explored. A large number of antlion species have been described from around world; still there is a lot of confusion in their taxonomy. In Pakistan, antlion fauna is not fully explored prior to this study only 28 species have been reported so far from the country. This study steps forward in exploration of Pakistan’s rich antlion faunal diversity. Still lot of it needs to be explored. This study improved the species number of Pakistan to 44. Which blong to 2 subfamilies 8 tribes and 20 genera. Subfamily Palparinae of Pakistan is represented with five genera and eight species. Species Palparellus astutus and Palpares rajasthanicus first time reportd from Pakistan. While Palpares trichogaster Navas, 1913 reinstate as valid species also P. cephalotes by Iqbal and Yousuf (1997) identified as junior synonym for the species. Genus Stenares Hagen, 1866 two species recorded Stenares hyaena clarus Navas, 1932 and Stenares improbus (Walker, 1853). Species S. improbus first time recorded from Pakistan. Author was not able to collect any specimen of genus Tomatares from Pakistan. Subfamily Myrmeleontinae is represented by seven tribes, eighteen genera and thirty five species. Tribe Acanthaclisini Navas, 1911 constitutes two genera Acanthaclisis and Centroclisis. Acanthaclisis obscura Hölzel, 1972 first time reported form Pakistan. Genus Centroclisis is represented by two species with one new species refered as Centroclisis thari.Tribe Dendroleontini Banks, 1912 is represented by one species Epacanthaclisis alaica first time recorded from Pakistan. In Tribe Nemoleontini Banks, 1911 represented by first time reported species Pseudoformicaleo gracilis. Tribe Neuroleontini Navas, 1912 with seven genera; genus Delfimeus Navas, 1912 monotypic Delfimeus intricatus. Genus Distoleon Banks, 1910 with four species Distoleon nefandus, Distoleon cyrus, Distoleon obducens and Distoleon tesselatus all four first reported from Pakistan Genus Ganguilus Navas, 1912 one species Ganguilus pulchellus (Banks, 1911). Genus Indoleon Banks (1913) presentdd with one species Indoleon vartianorum (Hölzel, 1972). Genus Neuroleon Navas, 1909 four species Neuroleon tenellus, Neuroleon (Ganussa) leptaleus, Neuroleon egenus and Neuroleon asiricus all first time reported. Myrmecaelurini Esben-Petersen, 1918 having six genera: Genus Gepus Navas, 1912 with one species Gepus invisus Navas, 1912 first time recorded from Pakistan. Genus Iranoleon, Hölzel, 1968 monotypic Iranoleon vartianae Hölzel, 1968. Genus Myrmecaelurus Costa, 1855 having four species. Myrmecaelurus laetus, Myrmecaelurus paghmanus and Myrmecaelurus (Nohoveus) vanharteni first time reported from Pakistan. Genus Solter with two species and Solter Solter felderi first time reported from Pakistan. Tribe Nesoleontini Markl, 1954 with three genera: Genus Cueta having five specie. While four species Cueta striata, Cueta thaliae, Cueta minervae and Cueta plexiformia first time recorded from Pakistan. Tribe Myrmeleontini only genus Myrmeleon with six species. Five species Myrmeleon trivialis,Myrmeleon bore, Myrmeleon hyalinus, Myrmeleon noacki and Myrmeleon inconspicuous are first time reported from Pakistan. Species were identified morphologically and 703 specimens were selected for DNA barcoding. Maximum 10 specimens from each species were selected. The Barcode Index Number (BIN) analysis using BOLD Systems Version 3.0 assigned 40 BINs to total 359 Antlion DNA sequences recorded for DNA barcode region. To find maximum distance distribution in species Barcode Gape Analysis and Automated Barcode Gap Analysis was performed using BOLD and online resource respectively. Results showed that maximum intera specific distance was 0 to 2.69%. While comparison with other countries was not possible as there was no data available for same species from other countries. This may be due to non processing of specimens from other countries or species endemism. The analysis showed that no species have been already sequenced and added to BOLD. So it has been added first time at least for local fauna. Value of maximum interspecific distance was less than nearresrt neighbor distance. Distance analysis of the barcode sequences showed an interspecific.divergence of at least 2%. Nearest neighbor joining (NNJ) tree also revealed that close taxa were grouped together more precisely than distant species. An online DNA based reference library was created, for species collected in the present study, on the Barcode of Life Data Systems (http://www.boldsystems.org). Results revealed that antlions exist throughout Pakistan in all geographical and environmental conditions and were not restricted to dry hot and arid areas. They are much abundant in undisturbed and non polluted areas. Collection and survey records revealed that in intensive crop zones due to habitat loss and higher chemical pollution antlion population has almost been eliminated.