Cytochrome C Oxidase Subunit Genes Research Articles

Preliminary study on buffy coat smear and molecular detection of microfilaria in domestic chickens (Gallus gallus domesticus) raised in Southern Thailand

Background and Aim: Filarial nematode typically produces a larval stage (microfilariae) in the bloodstream of vertebrate hosts, where microfilariae reside in the blood or subcutaneous tissues. Filarial nematodes cause human diseases, such as river blindness and elephantiasis, which are widely studied. However, in avian species, they are overlooked because they are nonpathogenic. In Thailand, microfilaria can be found in wild birds and domestic chickens. Recently, an increase in the number of blood samples submitted to veterinary diagnostic laboratories may have increased the number of microfilariae. Therefore, knowledge about filarial species and reliable detection methods are important. Therefore, this study aimed to investigate the efficacy of buffy coat smear and polymerase chain reaction (PCR)-based methods for the detection of microfilaria in domestic chickens. In addition, parasites were identified using the sequence of the cytochrome c oxidase subunit 1 (COX1) gene. Materials and Methods: Giemsa-stained buffy coat smears from a previous study were reanalyzed. These available buffy coat smears were prepared from 55 domestic chickens raised as backyard free-ranging in Southern Thailand. Fifty-seven frozen genomic DNA extracted from chicken blood were used to detect the presence of the COX1 gene in Onchocercidae nematodes. The nested PCR protocol for amplification of the OnchoCOI_ R2-OnchoCOI_ R2 fragment of the COX1 gene was applied from a previous report. Sequences of COX1 were analyzed to identify Onchocercidae nematodes and if they were single or mixed infections. We constructed Bayesian phylogenetics to identify parasites and assessment of the relationship between filarial nematodes in avian species and other vertebrate hosts. Results: Buffy coat smears from 15 samples revealed microfilaria. Of these 15 samples, only eight were positive for COX1 nested-PCR amplification. The other two buffy coat-negative samples were also positive for nested-PCR. Sequencing of these 11 nested PCR-positive samples revealed that almost all of them were Onchocercidae nematodes. Bayesian phylogenetic analysis showed that chicken Onchocercidae spp. were grouped with other avian filarial nematodes. However, all chickens Onchocercidae spp. showed a double peak in the sequencing chromatogram, indicating mixed filarial infection (species or haplotypes). Therefore, no chicken Onchocercidae sequence was deposited on National Center for Biotechnology Information, GenBank. Conclusion: Giemsa-stained buffy coat smear was a reliable method for the detection of chicken microfilaria in routine veterinary diagnostic laboratories. Development of a new PCR-based method is necessary. This method may provide greater sensitivity and specificity of detection. In addition, the PCR method allowed us to access the genetic characteristics of nematodes, which helped us maximize our knowledge of nematodes. Further investigations, such as the pathogenicity of filarial nematodes in chickens and their potential vectors, are required. Keywords: Buffy coat smear, Chickens, Cytochrome c oxidase I, Microfilaria, Nested-polymerase chain reaction.

In Silico Primer Design for geographical detection of Apis florea using Cytochrome c oxidase subunit 1 (COX1) gene

The yellow dwarf honey bee, Apis florea are well distributed in South Asia to South East Asia. This species is expanded and introduce area from their original distribution. However, the distribution of this honey bee in Indonesia is unexplored. The cytochrome c oxidase subunit 1 (COX1) gene are success to detect original geographic of introduce A. florea found in Egypt. The A. florea specific primer of COX1 gene are needed to produce the molecular marker for geographical origin detection. Thus, this study aims to in silico design the COX1 gene primer of A. florea using Primer3 and Primer-BLAST. This study results the best candidate primer of Af6_COX1_F and Af6_COX1_R primers that start from 212 to 983 regarding the A. florea COX1 gene (NC_021401) and produce 772 bp of amplicon. The melting temperature of forward primer are 54. 63 and 55.58°C, GC content of 40 and 45%, GC clamp of 3 and 2 for forward and reverse primer, respectively. There are no secondary structure of those primer and 100% homolog with A. florea COX1 sequences, thus those primer is potentially as the geographical origin marker of A. florea.

Classical morphology and DNA barcoding based identification of freshwater ectoparasite, Argulus foliaceus in rohu Labeo rohita

DNA barcoding coupled with classical morphological description have become promising approach for the species levelidentification. In the present study, 278 numbers of rohu Labeo rohita were screened, for Argulus infestation, amongwhich 167 fish (60.07%) were found infested with Argulus. Morphological attributes of the parasites were studied by lightmicroscope (LM) and Scanning Electron Microscope (SEM) which showed dorsoventrally flattened body comprising ofhead, thorax, abdomen, a pair of large compound eyes, suctorial organs with sclerotised support structures and two openingsof spermatheca at posterior end. Posterior incision of abdomen do not reach the mid-line and the cephalo-thoracic carapacehave not extended beyond the beginning of abdomen. Further validation was done using molecular tools for accurateparasite identification. The pair-wise genetic distance value, using Kimura-2 parameter showed a species level variationof 0.001 (1%) with that of A. foliaceus, while 0.083 and 0.052 (i.e. more than 2%) with that of A. indicus and A. japonicusrespectively. Phylogenetic tree generated using Neighbour-Joining (NJ) and Maximum-Likelihood (ML) methods, withKimura-2 parameter were also in agreement with pairwise distance values. The mitochondrial cytochrome c oxidase subunit1 (COI) sequences of A. foliaceus formed one cluster with the present studied samples, whereas sequences of A. japonicusformed a sister group. Integrating morphological and rapid DNA barcoding tools, the species was delineated as A. foliaceus. Keywords: Argulus foliaceus, COI gene, DNA barcoding, Labeo rohita, Morphology

Genetic Variation of Taenia Saginata Cyst Isolates from Iraq Based on Mitochondrial COX1 Sequences.

The zoonotic parasite disease of economic and public health relevance is bovine cysticercosis, resulting from the larval stage of Taenia saginata. The presented research aims to identify intraspecific variation in T. saginata isolated from cattle in Iraq's Sulaymaniyah province using the mitochondrial cytochrome c oxidase subunit 1 (COX1) gene. Sequence analysis of the COX1 gene revealed that five distinct haplotypes were identified in 37 T. saginata specimens from Iraq. Four of the five T. saginata haplotypes may have been identified for the first time in the world. Phylogenetic research revealed that all T. saginata haplotypes had been clustered in a single clade, with Korean and Iranian isolates sharing a high degree of closeness. In addition, individual haplotypes related to COX1 had a pairwise evolutionary divergence of 0.005- 0.013, whereas the overall evolutionary divergence regarding all five haplotypes ranged between 0.000-0.018. It was concluded that added newly recorded data on T. saginata genetic variation could have substantial implications for taeniasis epidemiology and control.

Mitochondrial Cox-1 Barcoding For The Identification and Phylogenetic Analysis of Sarcophaga (Liosarcophaga) dux Recorded From Jeddah City of Saudi Arabia

This study was aimed to identify and phylogenetically analyze the sarcophagid flies from Jeddah (Saudi Arabia) on the basis of mitochondrial cytochrome-C-oxidase subunit-1 (COX-1) gene. Five flesh fly samples were collected from different locations of Jeddah region. All the samples were first morphologically identified as Sarcophaga dux and confirmed on the basis of COX-1 gene amplification and sequencing techniques using universal primers LCO1490 and HCO2198. For phylogenetic analysis, 5 sequences were checked by COX-1 service on barcode of life data system (BOLD) database. To determine the phylogenetic relationships and genetic variability between 5 collected and BOLD samples, large Neighbor-joining tree comprising of the collected samples and all the available S. dux BOLD samples were constructed. The 32 most related BOLD sequences were used to construct maximum likelihood (ML) and Neighbor-joining trees. The phylogenetic analysis of COX-1 gene sequences revealed significant match with S. dux sequences from BOLD database: GQ254446 (Australia), MG780139 (India), AY879255 (China), JN869983 (Malaysia) and KC249664 (Egypt). This is the first report of the flesh fly species recorded in the Western regions of Saudi Arabia. Results indicated that the genetic diversity and phylogenetic relationship analysis of the collected S. dux samples have an important role in understanding the evolutionary processes of Sarcophaga cryptic species in Western regions of Saudi Arabia.

First report of molecular and phylogenetic analysis of Physaloptera praeputialis in naturally infected stray cats from India.

Nematodes of the genus Physaloptera are globally distributed and infect a multitude of hosts. Their life cycle involves orthopterans and coleopterans as intermediate hosts. The morphological characters alone are inadequate to detect and differentiate Physaloptera spp. from its congeners. Moreover, molecular studies are limited to compare them precisely. The present communication reports the first molecular phylogenetic characterization of feline Physaloptera spp. from India based on mitochondrial cytochrome c oxidase subunit 1 (COX1) and small subunit ribosomal DNA (18S rDNA). The nematodes were first isolated from the stomach of adult stray cats during necropsy examination. Based on the gross and microscopic characters, the worms were identified as P. praeputialis. Morphological identification was further confirmed through PCR targeting the barcode region of the mitochondrial cytochrome c oxidase subunit I (MT-COI) gene, using nematode-specific primers cocktail followed by species specific primers targeting partial COX1 and 18S rRNA genes. Generated sequences were submitted in NCBI GenBank (MW517846, MW410927, MW411349), and phylogenetic trees were constructed using the maximum likelihood method. When compared with other sequences of Physaloptera species across the globe, the present isolates showed 85.6-97.7% and 97.3-99% nucleotide homology based on COX1 and 18S rRNA gene, respectively. BLASTn analysis revealed a strong identity to other Physaloptera spp., and the phylogenetic tree placed all Physaloptera spp. in the same cluster. This study again indicates the usefulness of molecular techniques to substantiate the identity of species that may lack adequate descriptions and impart new insight for the potentially overlooked significance of P. praeputialis infections in felines.

Detection of novel mitochondrial mutations in cytochrome C oxidase subunit 1 (COX1) in patients with familial adenomatous polyposis (FAP).

Familial adenomatous polyposis (FAP) is an Autosomal dominant inherited disorder and a rare form‌ of colorectal cancer (CRC) that is characterized by the development of hundreds to thousands of adenomas in the rectum and colon. Mostly, cancers develop after the advent of the polyps. It appears in both sexes evenly, and the occurrence of the disease is in the second decade of life. Mitochondrial genome mutations have been reported with a variety of Tumors, but the precise role of these mutations in the pathogenicity and tumor progression is not exactly clear. Cytochrome c oxidase subunit I (COX1) is the terminal enzyme of the mitochondrial respiratory chain. The present study aims at assessing the occurrence of mtDNA mutations in COX1 gene in FAP patients and attempts to find out the cause and effect relationship between mitochondrial mutations and tumor progression. In this study, 56 FAP patients were investigated for the presence of the mutations in mitochondrial COX1 coding gene by PCR and sequencing analysis. All sequences that differed from the revised Cambridge Reference Sequence (rCRS) were classified as missense/ nonsense or silent mutations. Functional genomic studies using Bio-informatics tools were performed on the founded mutations to understand the downstream alterations in structure and function of protein. We identified 38 changes in the COX1 gene in patients with FAP symptoms. Most of them were heteroplasmic changes of missense type (25/38). Tree of the changes (G6145A, C6988A, and T7306G) were nonsense mutations and had not been reported in the literature before. Our results of bioinformatics predictions showed that the identified mutations can affect mitochondrial functions, especially if the conservative domain of the protein is concerned. Our findings indicate a high frequency of mtDNA mutations in all of the FAP cases compared to matched controls. These data significantly enhance our understanding of how such mutations contribute to cancer pathologies and develop the cancer treatment methods by new diagnostic biomarkers, and new drugs for gene therapy.

Analysis of the cytochrome c oxidase subunit 1 (COX1) gene reveals the unique evolution of the giant panda

As the rate-limiting enzyme of the mitochondrial respiratory chain, cytochrome c oxidase (COX) plays a crucial role in biological metabolism. “Living fossil” giant panda (Ailuropoda melanoleuca) is well-known for its special bamboo diet. In an effort to explore functional variation of COX1 in the energy metabolism behind giant panda's low-energy bamboo diet, we looked at genetic variation of COX1 gene in giant panda, and tested for its selection effect. In 1545 base pairs of the gene from 15 samples, 9 positions were variable and 1 mutation leaded to an amino acid sequence change. COX1 gene produces six haplotypes, nucleotide (pi), haplotype diversity (Hd). In addition, the average number of nucleotide differences (k) is 0.001629±0.001036, 0.8083±0.0694 and 2.517, respectively. Also, dN/dS ratio is significantly below 1. These results indicated that giant panda had a low population genetic diversity, and an obvious purifying selection of the COX1 gene which reduces synthesis of ATP determines giant panda's low-energy bamboo diet. Phylogenetic trees based on the COX1 gene were constructed to demonstrate that giant panda is the sister group of other Ursidae.

Utility of COX1 phylogenetics to differentiate between locally acquired and imported Plasmodium knowlesi infections in Singapore.

Although there have been several phylogenetic studies on Plasmodium knowlesi (P. knowlesi), only cytochrome c oxidase subunit 1 (COX1) gene analysis has shown some geographical differentiation between the isolates of different countries. Phylogenetic analysis of locally acquired P. knowlesi infections, based on circumsporozoite, small subunit ribosomal ribonucleic acid (SSU rRNA), merozoite surface protein 1 and COX1 gene targets, was performed. The results were compared with the published sequences of regional isolates from Malaysia and Thailand. Phylogenetic analysis of the circumsporozoite, SSU rRNA and merozoite surface protein 1 gene sequences for regional P. knowlesi isolates showed no obvious differentiation that could be attributed to their geographical origin. However, COX1 gene analysis showed that it was possible to differentiate between Singapore-acquired P. knowlesi infections and P. knowlesi infections from Peninsular Malaysia and Sarawak, Borneo, Malaysia. The ability to differentiate between locally acquired P. knowlesi infections and imported P. knowlesi infections has important utility for the monitoring of P. knowlesi malaria control programmes in Singapore.

Sequencing of COI gene in four rodent pests for species identification

AbstractAmong mammals, rodents are the most important group since they influence the crop production and protection. Species identification of rodents on morphometric parameters is cumbersome and often misleading. However, DNA based identification would enable accurate species identification. For this purpose, we examined 650 bp of Cytochrome c oxidase subunit1 (COI) in four rodent pest species namely

Haplotype Diversity of COI Gene of Hylarana chalconota Species Found at State University of Malang

Hylarana chalconota is a cryptic species of frog endemic to Java Island . This species is small with long legs, and brown skin. The Snout-Vent Length (SVL) ranges between 30-40 mm for male and 45-65 mm for female. Reports the existence of this species in State University of Malang, which was not found in 1995. To reveal the haplotype diversity of COI gene in this species, we analyzed Cytochrome-c oxidase subunit-1 (COI) sequences of both samples. The both samples had 604 bp and 574 bp fragment length, respectively. These fragments showed polymorphism; with mutation position in sites 104, 105, and 124. Based on this result, we suggest that the two samples share a different haplotypes, proposed as UM1 and UM2

Chromosome Conformation Capture of All 13 Genomic Loci in the Transcriptional Regulation of the Multisubunit Bigenomic Cytochrome c Oxidase in Neurons

Cytochrome c oxidase (COX) is the terminal enzyme of the electron transport chain composed of 13 subunits; three are mitochondria-encoded, and 10 are nucleus-inscribed on nine different chromosomes within the mammalian genome. The transcriptional regulation of such a multisubunit, multichromosomal, and bigenomic enzyme is mechanistically challenging. Transcription factories have been proposed as one mechanism by which genes from different genomic loci congregate to transcribe functionally related genes, and chromosome conformation capture (3C) is a means by which such interactions can be revealed. Thus far, however, only loci from the same chromosome or at most two chromosomes have been co-localized by 3C. The present study used 3C to test our hypothesis that not only the 10 genomic loci from nine chromosomes encoding the 10 nuclear subunits of COX, but also genes from three chromosomes encoding mitochondrial transcription factors A and B (Tfam, Tfb1m, and Tfb2m) critical for the transcription of the three mitochondria-encoded COX subunit genes all occupy common intranuclear sites in the murine neuronal nuclei. The pairing of various COX subunit genes and Tf genes indicates that interactions are present among all of them. On the other hand, genes for a non-mitochondrial protein (calreticulin) as well as a mitochondrial enzyme (citrate synthase) did not interact with COX genes. Furthermore, interactions between COX subunit and Tf genes were up-regulated by depolarizing stimulation and down-regulated by impulse blockade in primary neurons. Thus, a viable mechanism is in place for a synchronized, coordinated transcriptional regulation of this multisubunit, bigenomic COX enzyme in neurons.

Coupling of Energy Metabolism and Synaptic Transmission at the Transcriptional Level: Role of Nuclear Respiratory Factor 1 in Regulating both CytochromecOxidase and NMDA Glutamate Receptor Subunit Genes

Neuronal activity and energy metabolism are tightly coupled processes. Regions high in neuronal activity, especially of the glutamatergic type, have high levels of cytochrome c oxidase (COX). Perturbations in neuronal activity affect the expressions of COX and glutamatergic NMDA receptor subunit 1 (NR1). The present study sought to test our hypothesis that the coupling extends to the transcriptional level, whereby NR1 and possibly other NR subunits and COX are coregulated by the same transcription factor, nuclear respiratory factor 1 (NRF-1), which regulates all COX subunit genes. By means of multiple approaches, including in silico analysis, electrophoretic mobility shift and supershift assays, in vivo chromatin immunoprecipitation, promoter mutations, and real-time quantitative PCR, NRF-1 was found to functionally bind to the promoters of Grin 1 (NR1), Grin 2b (NR2b) and COX subunit genes, but not of Grin2a and Grin3a genes. These transcripts were upregulated by KCl and downregulated by tetrodotoxin (TTX) in cultured primary neurons. However, silencing of NRF-1 with small interference RNA blocked the upregulation of Grin1, Grin2b, and COX induced by KCl, and overexpression of NRF-1 rescued these transcripts that were suppressed by TTX. NRF-1 binding sites on Grin1 and Grin2b genes are also highly conserved among mice, rats, and humans. Thus, NRF-1 is an essential transcription factor critical in the coregulation of NR1, NR2b, and COX, and coupling exists at the transcriptional level to ensure coordinated expressions of proteins important for synaptic transmission and energy metabolism.

Is nuclear respiratory factor 2 a master transcriptional coordinator for all ten nuclear-encoded cytochrome c oxidase subunits in neurons?

Cytochrome c oxidase (COX), the terminal enzyme of the mitochondrial electron transport chain, is a multi-subunit, bigenomically encoded inner mitochondrial membrane protein. Of the thirteen subunits, three are encoded in the mitochondrial genome and ten others are encoded in the nuclear genome. Transcriptional coordination of nuclear-encoded COX subunit genes is likely accomplished by transcription factors responding to upstream signals. Previous studies have found that nuclear-encoded COX subunit genes are under the control of specific transcription factors, such as nuclear respiratory factor 2 (NRF-2). However, it is not known if a single transcription factor binds to all ten of COX subunit promoters. In the current study, we identified in silico putative NRF-2 binding sites on all ten nuclear-encoded COX gene promoters in the rat genome. Chromatin immunoprecipitation assay showed that NRF-2 bound in vivo to six of the ten nuclear-encoded COX subunit promoters. Electrophoretic mobility supershift assays demonstrated binding of NRF-2 to the other four subunits, and promoter mutation study confirmed the functionality of these NRF-2 binding sites. Finally, transfection of dominant-negative constructs of NRF-2 proteins caused a significant reduction of COX expression. We conclude that NRF-2 is an important mediator of coordinated regulation of all ten nuclear-encoded COX subunit genes in neurons.

Expression of cytochrome c oxidase subunit 1 gene in the brain at an early stage in the termination of pupal diapause in the sweet potato hornworm, Agrius convolvuli

Suppression-subtractive hybridization was used to isolate cDNAs specifically expressed in the brain at the termination of pupal diapause in Agrius convolvuli. One of the isolated clones shows similarity to the cytochrome c oxidase subunit 1 (COX1) gene. The full-length cDNA was obtained from brain mRNA by rapid amplification of cDNA ends (RACE). The insert is 1.65 kb in length and has an open reading frame of 1.46 kb which encodes a putative protein of 486 amino acid residues. RT-PCR reveals that the mRNA increases dramatically at an early stage of diapause termination. Activity of cytochrome c oxidase in the brain also increases at the same time. The up-regulation of this gene suggests that expression of the COX1 gene and ATP synthesis are initiated in the brain in association with diapause termination.

Regulation of cytochrome c oxidase subunit 1 (COX1) expression in Cryptococcus neoformans by temperature and host environment.

In the study of differential gene expression of Cryptococcus neoformans, a transcript of COX1 (cytochrome oxidase c subunit 1) was identified in a serotype A strain. The transcript was upregulated at 37 degrees C compared to 30 degrees C and expressed by yeasts infecting the central nervous system. Northern analysis of COX1 from the serotype A strain revealed two polycistronic transcripts, a temperature-upregulated 2.3 kb transcript and a 1.9 kb transcript that was not affected by temperature. In contrast, COX1 in a serotype D strain showed only a 1.9 kb polycistronic transcript plus a 1.6 kb monocistronic message, and temperature had no effect on the transcripts. The sequence of COX1 revealed similar coding regions between the two strains, but the serotype D strain had five introns whereas no introns were found in the serotype A strain. The serotype D strain had reduced growth rates compared to the serotype A strain at 37 degrees C, but in an AD hybrid strain the serotype D COX1 gene could support efficient high temperature growth. These studies have revealed mitochondrial molecular differences between serotype A and D strains which show evolutionary divergence. It will be important to determine whether differences in mitochondrial structure and function can influence cryptococcosis.

Cytochrome c oxidase deficiency.

Cytochrome c oxidase (COX) is the terminal enzyme of the mitochondrial respiratory chain, catalyzing the transfer of electrons from reduced cytochrome c to molecular oxygen. It is composed of 13 structural subunits, three of which are encoded in mtDNA and form the catalytic core of the enzyme. In addition to these structural subunits, a large number of accessory factors are necessary for the assembly and maintenance of the active holoenzyme complex. Most isolated COX deficiencies are inherited as autosomal recessive disorders; mutations in the mtDNA-encoded COX subunit genes are relatively rare. These mutations are associated with a wide spectrum of clinical phenotypes ranging from isolated myopathy to multisystem disease, with onset from late childhood to adulthood. Autosomal recessive COX deficiencies generally have a very early age of onset and a fatal outcome. Several clinical presentations have been described including Leigh Syndrome, hypertrophic cardiomyopathy and myopathy, and fatal infantile lactic acidosis. Surprisingly, mutations in the nuclear-encoded structural COX subunits have not been found in association with any of these phenotypes. Mutations have, however, been identified in several COX assembly factors: SURF1 (Leigh Syndrome), SCO2 (hypertrophic cardiomyopathy), SCO1 (hepatic failure, ketoacidotic coma), and COX10 (encephalopathy, tubulopathy). As all of these assembly factors are ubiquitously expressed, the molecular basis for the different clinical presentations remains unexplained. Although the genetic defects in the majority of patients with COX deficiency are unknown, it is likely that most will be solved in the near future using functional complementation techniques.

A systematic mutation screen of 10 nuclear and 25 mitochondrial candidate genes in 21 patients with cytochrome c oxidase (COX) deficiency shows tRNA(Ser)(UCN) mutations in a subgroup with syndromal encephalopathy.

COX deficiency is believed to be the most common defect in neonates and infants with mitochondrial diseases. To explore the causes of this group of disorders, we examined 25 mitochondrial...

Mitochondrial DNA deletions and cytochrome c oxidase deficiency in muscle fibres

We have studied cytochrome c oxidase (COX) deficient muscle fibre segments in 6 patients with mitochondrial myopathy and deletions of mitochondrial DNA (mtDNA). The distribution of transcripts of normal and mutated mtDNA in skeletal muscle sections was studied by in situ hybridization. The results were compared with the enzyme histochemical activity of COX and the immunohistochemical distribution of mtDNA encoded and nuclear DNA encoded subunits of COX. In all cases a proportion of the muscle fibres (< 1–30% of the fibres in cross-sections) and had low COX activity and high activity of succinate dehydrogenase (COX deficient muscle fibres). Transcripts of normal and deleted mtDNA showed the same distribution within the tissue as the corresponding mtDNA, indicating that the deleted mtDNA is transcribed. The COX deficient muscle fibres showed accumulation of transcripts of deleted mtDNA, which had a similar distribution as the accumulated mitochondria within these fibres. With few exceptions, there was a low level of transcripts of normal mtDNA in these COX deficient fibres. Immunohistochemical analysis revealed low levels of immunoreactive material using antiserum to the mtDNA encoded subunits II/III as well as the nuclear DNA encoded subunit IV of COX in all COX deficient muscle fibres. The fraction of deleted mtDNA in muscle ranged from 43 to 87%. There was no correlation between the proportion of COX deficient muscle fibres and the fraction of deleted mtDNA. In 2 cases the deletion did not involve any COX gene. One of these cases had 87% deleted mtDNA but less than 1% COX deficient muscle fibres. This case had several muscle fibres with accumulation of transcripts of deleted mtDNA concomitant with normal COX activity. The other of these 2 cases had 10% COX deficient muscle fibres. Our results show that the proportion of COX deficient muscle fibres is not correlated to the fraction of deleted mtDNA in muscle tissue and that a high proportion of COX deficient fibres may be present even if COX subunit genes are not affected by the deletion. Reduced enzyme histochemical activity of COX in muscle fibre segments is correlated to low levels of normal mtDNA in these fibre segments.