© JAPI • August 2012 • VOL. 60
*Consultant Biochemist, Head, Dept. of Laboratory Medicine, Jt.
Director Research, **Molecular Biologist, ***Research Fellow, PD Hinduja
National Hospital and Medical Research Centre, Lalita girdhar
Building, Veer savarkar Marg, Mumbai 400016, Maharashtra
Received: 03.12.2010; Revised: 09.05.2011; Accepted: 25.05.2011
transport of chloride ions across the apical membranes of
ystic Fibrosis (CF) is one of the most lethal, autosomal,
recessive monogenic disorder caused due to an abnormal
the sweat glands in CF are relatively impermeable to chloride
ions resulting in the increased concentration of chloride in the
sweat reaching the skin surface. to maintain electroneutrality,
the reabsorption of sodium ions by sweat glands is also reduced,
and therefore an increased concentration of sodium also has been
observed in the sweat.1 thus, a defect in the transport of chloride
and sodium ions is characteristic of tissues affected in CF.
However, normal chloride and sodium transport has been
observed in some CF cases, indicating the contribution of other
pathophysiologic processes. A reduced secretion of bicarbonate
ions across the epithelial cells has been reported in some CF cases.
Bicarbonate, the body’s major buffer, maintains the alkaline pH
in the epithelia, and a reduced bicarbonate secretion, creates an
acidic environment leading to the precipitation of mucins and
plugging of ductal systems.2
the salt or ion movements establish the osmotic driving
force for water movement within the tissues. the abnormal
ion transport observed in CF, disturbs the hydration state of
secretions in the epithelial lumen, due to which the secretions
from various glands become dehydrated, thick and sticky,
instead of being watery and free-flowing. this eventually
leads to plugging of mucous secretions in the ducts of exocrine
glands of respiratory, pancreatic, intestinal, biliary and male
CFTR Gene Mutations
the gene responsible for CF is called the Cystic Fibrosis
transmembrane Conductance Regulator (CFtR) gene. It encodes
the amino acid product termed the CFtR protein, functioning as
a chloride channel. More than 1000 population-specific mutations
have been reported in the CFtR gene.4 these mutations may be
mild or severe resulting in partial or complete non-functioning
of CFtR chloride channel respectively.
One of the most severe, prevalent and predominant mutation
in all the populations is the deletion of phenylalanine at amino
acid position 508 (DF508) of the CFtR protein.5
CF was considered to be extremely rare in India. the
disease was first described in an Indian patient in 1968 from
the Post graduate Institute of Medical Education and Research,
Chandigarh. since then, the published data on Indian CF
patients, however, has been very limited. the precise incidence
of CF in India is still not known; and the information on CF
mutations is also very scarce.
genetic studies on CF is being carried out at a few research
centres in India including P. D. Hinduja National Hospital and
Medical Research Centre, Mumbai; All India Institute of Medical
sciences, New Delhi and the Post graduate Institute of Medical
Education and Research, Chandigarh.
The ΔF508 mutation has been reported to exhibit a frequency
of approximately 19 - 56% in the Indian CF cases,6-9 as compared
to 70% in the West. This low frequency may be indicative of the
differences in the relative frequencies of CFTR mutations in the
Indian CF cases, as compared to the western population.8
A screening study of the CFtR gene carried out on
Indian CF patients has reported the identification of 2 novel
mutations (3622Inst, 3601-20t→C/u) and 2 rare mutations
(3849+10KbC→t/u, R560H), along with DF508 mutation, thereby
anticipating a different spectrum of CF mutations in Indians.8
A similar data has been observed during our study carried
out at P. D. Hinduja National Hospital and Medical Research
Centre, wherein, we attempted at identifying and determining
the frequency of 6 of the most common mutations of the world
CF population including DF508, g542X, g551D, R553X, N1303K
and 621+1(g→t) mutations in 23 suspected Indian CF cases, by
multiplex ARMS-PCR technique.10 the DF508 mutation, during
this study, was observed to possess a frequency of 33%; however,
none of the other common mutations were identified.
We then screened the hot spot regions of the CFtR gene (exons
10 and 11) by single stranded Conformation Polymorphism /
Heteroduplex (SSCP/HD) technique in 139 clinically suspected
CF cases. This study, lead to the identification of a rare splice-site
mutation (1525-1(g→A) and a common polymorphism (M470V),
in addition to the DF508 mutation.10 the 1525-1(g→A) mutation
Cystic Fibrosis in India: A Systematic Review
Tester F Ashavaid*, Rani Raghavan**, Pradnya Dhairyawan***, Shweta Bhawalkar**
Objectives: CF, caused due to abnormal transport of chloride, sodium and bicarbonate ions across epithelial cell
membranes, is a multi-organ disorder. More than 1000 mutations causing CF, have been identified in the CFtR
gene, of which AF508 is the most severe, predominant mutation. However, data on CF in India is limited. Also,
facilities for CF diagnosis are not available at all diagnostic centres across India.
Results: AF508 mutation has been reported in 19 - 56% Indian patients. Also, the spectrum of mutations has been
anticipated to be different, due to the identification of a wide range of novel and rare mutations. In addition
to mutations, polymorphisms with clinical relevance and practical diagnostic value have also been identified.
Clinical profile in Indian patients was also observed to be different.
Conclusion: though, Cystic Fibrosis has always been considered to be a rare disease in India, we hope that the
identification of the wide range of mutations, leads us to the recognition of a probable increased incidence of CF in Indian
patients. And this would attract greater attention to the diagnosis of this disease, so that a clinically appropriate
assay can be developed for their detection as a preliminary test for CF diagnosis. the results observed during
the study can be a step forward in planning a molecular screening and providing appropriate genetic counseling
programs, which are lacking in our country at the moment.
40 © JAPI • August 2012 • VOL. 60
is predicted to code for a class II defective CFtR protein and
hence, classified as a severe pathogenic mutation.11 However,
the exact frequency and diagnostic value of this mutation in
Indians remains to be established with a larger study population.
We then screened the entire CFtR gene in 96 Indian CF
patients in a recent study, by the ssCP analysis, with the aim
of identifying all the known as well as unknown mutations.
During this study, a total of 14 mutations, including 09 novel
[-219insg, -117(g→C), 185+1(g→C), R59X, R75g, 405+1(g→C),
s169g, N187D, 3600+6(t→C)] and 05 rare, known (s13F, R75Q,
1525-1(g→A), ∆F508, Y569D) mutations were identified in 88
out of 96 CF patients. While, the DF508 mutation was found
to possess an allele frequency of approximately 53%; the
remaining mutations were observed to be rare, with most of
the mutations being detected in single CF cases (table 1). In
addition, 20 polymorphisms were also identified, of which 7
were novel polymorphisms [-500(A→g), 1342-15(g→t), 4096-
265insg, 4096-268inst, L188L, L167L, 622-92(C→A)] while the
remaining 13 [1525-61(A→g), M470V, 1898+152(t→A), t854t,
4005+121deltt, 4521(g→A), 1001+11(C→t), 5/6ttgA repeats,
622-100(A→g), 405+46(g→t), 297-67(A→C) and Poly t] were
known polymorphisms. Of these, some polymorphisms for e.g.,
M470V, poly t have been reported to be of clinical relevance and
might be of practical diagnostic value.12
Considering the identification of a high percentage of rare and
novel mutations, and the ethnic history of Indian population, it
may be speculated that the remaining uncharacterized mutations
might also not be prevalent mutations, and may prove to be
either private mutations or novel mutations. the characteristic
CFTR gene mutation distribution pattern in the Indian patients
and the mutational heterogeneity observed is in complete
agreement with the diverse heterogeneous ethnic origin of the
Clinical Presentations in Indian CF
the Indian CF patients mainly present with respiratory
and gastrointestinal problems associated with malnutrition.
Among these varied clinical symptoms, pulmonary involvement
has been observed to be the most predominant and severe CF
manifestation.13 this is also in accordance with the observation
made during our study, according to which almost 94% of the
Indian CF cases exhibit respiratory abnormalities (table 2).
Although majority of CF cases present during infancy and
childhood, a number of cases have been diagnosed in the
adulthood. During our study, the cases presenting pancreatic
abnormalities especially were observed, to possess a higher age
group, indicating that the damage of pancreas inutero occurs
progressively; and the patients can present with symptomatic
pancreatic abnormalities as the initial manifestations of CF in
According to the literature, CF cases with pancreatic
abnormalities exhibits the presence of ΔF508 mutation in the
homozygous or heterozygous state.15 However, in our study,
majority (>90%) of Indian CF cases with pancreatic abnormalities
did not exhibit the presence of ΔF508 mutation, probably
suggesting a different clinical profile in the Indian CF cases.10
Clinical severity of CF disease is thus a combination of CFtR
mutations influenced by other genetic and environmental
factors.16 Marked differences in the clinical severity were
observed among genetically identical homozygous ΔF508 cases
in the present study, indicating the modifying influence of other
genetic factors, such as the presence of a second mutation on the
same CFTR allele, attenuating the effect of ΔF508 mutation or the
effect of other non-genetic factors such as antibiotic treatment,
nutrition, etc. Environmental factors like pollution or tobacco
exposure may also play a role.
CF being a recessive disorder, the expression of the disease
requires the presence of mutations on both the CFTR alleles.
The heterozygous ΔF508 cases may therefore, harbor a second
unidentified mutation. However, a single CFTR mutation in
combination with specific alleles of other genes or an unfavorable
environment can also produce a CF phenotype.17
CF Diagnosis in India
Variability in the type and severity of CF clinical presentations
results in frequent errors of labeling CF with a different diagnosis.
the respiratory manifestations mimic those of bronchitis,
whooping cough, asthma and chronic lung diseases such as
immune deficiency disease, tuberculosis, fungal infections of
the lungs, bronchiectasis, etc. the gastrointestinal manifestations
mimic those of celiac disease, chronic diarrhoea and numerous
conditions associated with failure to thrive. thus diagnosis of
CF can be easily missed due to a low index of suspicion among
the Indians. Also variability in the severity and type of clinical
manifestations often lead to delayed diagnosis. thus, CF may be
far more common in people of Indian origin than was previously
thought; but is under diagnosed or missed in majority of cases.
the diagnosis of CF is suspected by the presence of a typical
CF phenotype and/or family history of CF. CF being a genetic
disease, a positive family history should be an important factor in
suspecting the disease. However, in view of the recessive mode
of inheritance of CF and the present trend towards small families,
Table 1 : Fourteen CFTR mutations identified in 96 Indian
CF patients (*Novel mutations)
Mutations Location in
Table 2 : Clinical presentations observed in Indian CF
Family history of CF
*Majority of the CF patients exhibited multi-organ symptoms
© JAPI • August 2012 • VOL. 60
the majority of CF cases appear only in a single member of the
family, as a result of which the family history does not provide
much diagnostic assistance.18
The suspected CF cases are confirmed by the demonstration
of a high sweat chloride (>60mmol/L) concentration. However,
sweat testing facilities are not available in most centres in India.
the high initial and recurring cost of this test probably makes
it less suitable for use in all centres. this poor availability of
facilities for CF diagnosis may also be responsible for the under
diagnosis and low incidence of CF in India.
A CF patient exhibiting typical CF clinical presentations along
with a homozygous or heterozygous ΔF508 mutation may have a
normal sweat electrolyte concentration, if a second ameliorating
or neutralizing mutation is present elsewhere on the CFtR allele.
this suggests that elevated sweat electrolytes may be diagnostic
of CF, but a negative sweat test does not completely exclude the
possibility of CF.19
According to the literature,20 it is unusual for a child to have a
sweat chloride concentration greater than 30-40mmol/L; and the
sweat chloride concentrations increase with age after puberty.
Thus, in order to avoid any misdiagnosis, 2 different upper limits
(30-40mmol/L in children and the traditionally used 60mmol/L
in adults) of sweat chloride concentration may be implemented.
the suspected CF cases with a borderline sweat chloride
concentration (40-60 mmol/L) presents a diagnostic challenge,
as the diagnosis of CF can neither be confirmed nor excluded.
In such circumstances, mutation analysis or DNA testing can
substitute for the sweat test. the presence of mutations in the
CFtR gene can predict with a high degree of certainty that an
individual has CF.21 Also, in some suspected CF cases, especially
in infants, the adequate collection of sweat sample becomes
difficult and mutation analysis can aid in CF diagnosis, in such
Mutation analysis in several populations consists of the
screening of a panel of the most common CFtR mutations, so that
a mutation detection rate of greater than 90% can be achieved.5
The identification of the CF mutations commonly seen in any
given population is valuable when mutation analysis is used
as a ‘diagnostic test’ for CF. However, since the total number
of CF-causing mutations in the Indian patients is likely to be
very large, a DNA-based population screening in India will be
complicated; and an indirect genetic diagnosis like screening
the entire gene by SSCP, DGGE, DNA sequencing, etc. may be
necessary to characterize all the CFtR mutations in our patients.
though, Cystic Fibrosis has always been considered to be a
rare disease in India, we hope that the identification of the wide
range of novel and rare mutations, leads us to the recognition of
a probable increased incidence of CF in Indian patients. And this
would attract greater attention to the diagnosis of this disease,
so that a clinically appropriate assay can be developed for their
detection as a preliminary test for CF diagnosis.
Further functional studies will be required to determine
whether the molecular mechanisms involved in the base
pair substitutions reported in this study could lead to subtle
variations in levels of CFtR expression.
the results observed during the study can be a step forward
in planning a molecular screening and providing appropriate
genetic counseling programs, which are lacking in our country
at the moment.
Knowledge about all the mutations will be an essential part
in understanding the structure and function of the protein. In
conjunction with the development of the three-dimensional
structural analysis and tests for the biological activities of CFtR,
it will be possible in future, to define the role of all these residues
at the molecular level, and eventually, understand the function
of the CFtR protein.
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