AJCP / Meeting Abstracts
Detection of Putative Novel Mutation in Zp1 Gene
Causing Empty Follicle Syndrome
D. Upadhyay1, S. Anandt1, B. Peramo2, H. Al Safar3;
Genetics Department, Al Ain Fertility Center, Al Ain,
Abu Dhabi, United Arab Emirates, 2Medical Director,
Al Ain Fertility Center, Al Ain, Abu Dhabi, United Arab
Emirates, 3Center for Biotechnology, Khalifa University,
Abu Dhabi, Abu Dhabi, United Arab Emirates
Introduction/Objective: Mutations in the zona pellucida
glycoprotein genes have been reported to be associated
with empty follicle syndrome (EFS) and abnormal zona
pellucida (ZP). At our clinic, in a cohort of infertile pa-
tients, we found three cases with empty follicle syndrome
as a root cause of infertility. Further, we designed a study
to identify the disease-causing gene mutations in these
patients. At present, there are no citations or functional
evidence in ClinVar for NM_207341.3:c.1168del variant
indicating the involvement of this variant in empty fol-
licle syndrome. But it has been interpreted to be Likely
pathogenic causing female infertility due to zona pellu-
cida defect.
Methods/Case Report: In this prospective study con-
ducted at Al Ain Fertility Center (AAFC), U.A.E, we
performed genetic analysis on three patients (first-degree
sisters) having a mean age of 31.7 (2.52) years. Patient
indications were primary female infertility and IVF
failures due to no eggs present in the follicles when egg
retrieval was attempted. We characterized these three pa-
tients as suffering from empty follicle syndrome. Along
with the patient's sample, we also included their common
parent's sample for Whole Exome Sequencing and Sanger
sequencing, to rule out the inheritance pattern in the
family. The effects of this mutation were further charac-
terized through mRNA and protein studies.
Results (if a Case Study enter NA): N/A.
Conclusion:  To our knowledge, this is the first study in the
U.A.E. showing the familial novel variant in the ZP1 gene
associated with empty follicle syndrome. We identified a
novel mutation in these patients (NM_207341.3:c.1168del)
in the ZP1 gene in the homozygous state and the same
variant in the heterozygous state in the parents. Our find-
ings add to the existing mutational spectrum of the ZP1
gene associated with empty follicle syndrome and ab-
normal oocytes and provide new support for the genetic
diagnosis of female infertility.
The Assessment of the Diagnostic Accuracy of Antigen-
Detecting Rapid Diagnostic Test using Nasopharyngeal
Swab Specimen in Detecting SARS-CoV-2 among
Patients in a Private Hospital in Bulacan, Philippines
R.M. Magpoc1, M. Murzo1, A.P. Jayag1, S.P. Lee1,
A.M. Manguiat1, E.M. Oloteo1, A. Mendoza1,
S142 Am J Clin Pathol 2022;158:S30-S161
DOI: 10.1093/ajcp/aqac126
J.S. Hizon1, J.C. Macalalad1, R.G. Sison1, X.A. Nieto2;
1
Department of Medical Technology, University of Santo
Tomas Manila, Manila, Metro Manila, Philippines,
2
College of Science, Department of Mathematics and
Statistics, University of Santo Tomas Manila, Manila,
Metro Manila, Philippines
Introduction/Objective:  Continuous progression of
the COVID-19 outbreak prompted an expensive, lim-
ited, and slow-moving laboratory-based molecular
testing capacity of the gold standard, Real-time Reverse
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Transcription- Polymerase Chain Reaction (rRT-PCR),
in detecting SARS-CoV-2. The vital step in mitigation is
utilizing an accurate diagnostic detection assay to deploy
various preventive measures. Therefore, there is a need
to evaluate an accessible, fast, and inexpensive point-
of-care immunoassay, such as Antigen-detecting Rapid
Diagnostic Tests (Ag- RDTs). Thus, the study aimed to
assess the diagnostic accuracy of Ag-RDT in detecting
SARS-CoV-2.
Methods/Case Report: A non-experimental, quantita-
tive, and cross-sectional cohort study utilized 272 SARS-
CoV-2 Ag-RDT with rRT-PCR confirmation collected
from April 2021 to February 2022 in a private primary
hospital laboratory database in Bulacan, Philippines.
Diagnostic accuracy parameters of Ag-RDT included
in the study are sensitivity, specificity, positive predictive
value (PPV), and negative predictive value (NPV).
Results (if a Case Study enter NA):  The overall sensitivity
and specificity of Ag-RDT were 69.6% and 98.8%, re-
spectively. Additionally, it exhibited a PPV of 84.2% and
an NPV of 97.2%. With a hypothetical 1% prevalence,
PPV and NPV were 36.8% and 99.7%. Meanwhile, PPV
and NPV were 86.5% and 96.7% at 10% prevalence. High
specificity denotes that positive Ag-RDT test results can
be confidently considered in true SARS-CoV-2 infection,
while low sensitivity entails that a negative result must be
subjected to confirmatory tests. Furthermore, when prev-
alence increases, PPV increases while NPV decreases.
Conclusion: The study primarily encapsulated that the
Ag-RDT cannot replace rRT-PCR in detecting SARS-
CoV-2 infections. Despite its shortcomings, Ag-RDT is
still valuable in areas with moderate-to-high disease prev-
alence and with limited access to confirmatory test due to
its high predictive values and advantages. A larger number
of positive results of rRT-PCR, evaluation of factors cor-
relating to low sensitivity, and a prospective study are re-
commended for future studies.
Analysis of microRNAs by the Stem-Loop Real-Time
Quantitative Reverse Transcription Polymerase Chain
Reaction Using A Double Quencher Probe
A. Kikuchi1, A. Naruse1, K. Takagi2; 1Research
Institute for Medical Science, Daiyukai Health System,
© American Society for Clinical Pathology

Ichinomiya, Aichi, Japan, 2Department of Urology,
Daiyukai Daiichi Hospital, Ichinomiya, Aichi, Japan
Introduction/Objective:  We created a universal probe that
can be used with mature micro RNAs (miRNAs) using a
double quencher incorporating the ZEN quencher into the
hydrolysis probe required for miRNA analysis by the real-
time quantitative polymerase chain reaction (RT-qPCR)
using a stem-loop primer. The main purpose of this ap-
proach was to improve sensitivity and convenience.
Methods/Case Report:  We conducted studies on miR-21,
miR-30a and miR-200c. To assess the relationship be-
tween the sequence interval and reactivity between the
miRNA-specific sequence on the stem-loop primer and
detection probe, various sequence patterns were examined
using miRNA from K562 cells. The sensitivity and reac-
tivity of the designed primers and probes were examined
using each synthetic miRNA and transurethral resection
of bladder tumor (TURBT) samples. The synthetic RNA
was serially diluted in 10% increments. Twenty-seven
TURBT cases were examined. The prepared samples
were extracted with the FastGene RNA Basic Kit and
FastGene miRNA Enhancer. After extraction, cDNA
was prepared using FastGene Scriptase II. We performed
quantitative analysis by the RT-qPCR using a ZEN
double quencher probe. The samples were analyzed with
the Taq Man miRNA assay.
Results (if a Case Study enter NA): The probe and
miRNA-specific sequences of the stem-loop primer
showed better reactivity when the design contained less
space within the sequences. Serial dilution of the samples
using synthetic miRNA showed that linearity depended
on the addition amount, and occurred at a concentration
of log 7 or more. In the TURBT samples, the Δ of the
mean Cp value, and the correlation coefficients of miR-
21, miR-30a, and miR-200c, were 3.13 and 0.966; 1.10
and 0.991; and 0.39 and 0.997, respectively, compared
with the TaqMan miRNA assay.
Conclusion:  This demonstrates that our method has the
same or higher sensitivity than the TaqMan miRNA
assay. Overall, these findings suggest that the analysis
of miRNA with a stem-loop primer using the double
quencher probe is useful.
Clinico-Demographic Characteristics of Patients with
SARS-CoV-2 Antibodies: A Single Tertiary Center
Retrospective Study in Bohol, Philippines
A.L. Salillas, A.B. Clapis, E.G. Almocera, A.T. San Juan;
Pathology, Gov. Celestino Gallares Memorial Hospital,
Tagbilaran, Bohol, Philippines
Introduction/Objective: Coronavirus Disease 2019
(COVID-19) caused by SARS-CoV-2 continues to be
a global threat until the present. Most of the infected
© American Society for Clinical Pathology
AJCP  / Meeting Abstracts
patients are asymptomatic which can be a challenge in
controlling the spread of disease. Antibody detection
by chemiluminescence immunoassay (CLIA) is a good
complementary test to Real-Time Reverse Transcription
– Polymerase Chain Reaction (rRT-PCR) as an indirect
marker for disease.
Methods/Case Report:  This retrospective study assessed
the clinico-demographic characteristics of 184 SARS-
CoV-2 IgM and IgG antibody-reactive patients at Gov.
Celestino Gallares Memorial Hospital from July 1, 2020 –
May 31, 2021.
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Results (if a Case Study enter NA):  A total of 184 cases
were included in the study out of 4,138 patients tested
for anti-SARS-CoV-2 IgM and IgG antibodies by CLIA.
Within this population, 97 were outpatients and 87 were
inpatients. Majority of patients needing hospitalization
were aged ≥60-year-old males (33.33%) and that the prob-
ability of being admitted decreases with younger age.
Most of the outpatients who were reactive to antibodies
against SARS-CoV-2 were frontline workers compared to
non-frontline workers and non-workers (27.84% versus
1.03% and 19.59%). Most common comorbidities were
hypertension (24.46%), cardiovascular disease (15.76%),
diabetes mellitus (11.41%), and chronic kidney disease
(11.41%). All five mortalities were diagnosed hypertensive
who later developed critical disease.
Conclusion:  The continued use of antibody detection by
CLIA as a complementary test with rRT-PCR is still re-
commended not only for better detection of COVID-19
especially in asymptomatic patients but also to observe
changes in antibody titer especially in the advent of
COVID-19 vaccines.
Rapid Detection of Uropathogens and Antibiotic-resistant
Genes Using Open Array Multiplex PCR Technology
P. Neopane1, J. Nypaver1, S.S. Beqaj1, R. Shrestha2;
1
Molecular, Patients Choice Laboratories, Zionsville,
Indiana, United States, 2Patients Choice Laboratories,
Indianapolis, Indiana, United States
Introduction/Objective:  Introduction: Standard diag-
nostic practices for Urinary Tract Infection (UTI) take
2–3  days for pathogen identification and antibiotic sus-
ceptibility testing leading to delayed diagnosis and an in-
crease in healthcare expenditures. The rapid diagnostics
tool is highly desirable to identify the pathogen and its
antibiotic-resistant genes directly from the samples. Our
objective is to validate and evaluate the performance of
Open array multiplex PCR for pathogen identification
and antibiotic resistance genes.
Methods/Case Report: Method: Open array multi-
plex PCR method for 20 UTI pathogens was validated
using ATCC organisms and inter-laboratory corre-
lation using 50 urine samples. Briefly, samples were
Am J Clin Pathol 2022;158:S30-S161 S143
DOI: 10.1093/ajcp/aqac126