Identification of suspected paragonimiasis-endemic foci using a questionnaire and detection of Paragonimus ova using the Ziehl–Neelsen technique in Zamboanga Region, the Philippines

ABSTRACT

Improving paragonimiasis surveillance, which is crucial for disease control, requires adopting new tools and techniques useful in mapping endemic areas. This study aimed to (1) develop a questionnaire to identify suspected paragonimiasis-endemic foci, (2) describe the epidemiology of paragonimiasis, and (3) evaluate Ziehl–Nielsen Staining technique (ZNS) in detecting Paragonimus ova. The questionnaire, which municipal health officers filled out, was based on proposed site inclusion criteria utilized in the integrated tuberculosis (TB)-paragonimiasis surveillance and control project. Newly deployed medical technologists in Zamboanga Region underwent training, which included laboratory diagnosis of paragonimiasis using preserved and fresh specimens and an integrated tuberculosis-paragonimiasis survey in nine selected barangays (villages). Paragonimiasis cases were found in seven out of the nine barangays identified by the questionnaire. Of the 373 patients, three (0.80%) were TB-positive, and 29 (7.77%) were paragonimiasis-positive. The highest paragonimiasis prevalence (27%) was found in Barangay Libato. Ziehl–Neelsen Staining technique (ZNS) correctly detected 8 out of the 29 samples positive (sensitivity – 27.59%; 95% CI: 12.73–47.24%) and all the 334 samples negative (specificity – 100%; 95% CI: 98.90–100%) for Paragonimus ova. The questionnaire may be improved by refining the inclusion criteria. In paragonimiasis-endemic areas, the ZNS and the NaOH concentration technique may be used for detecting Paragonimus ova. Modifying the ZNS, for instance by including a concentration step, may improve its sensitivity. The model for the integrated capacity building of health workers and surveillance and research demonstrated in this project may contribute to improving surveillance and control of paragonimiasis and other neglected tropical diseases.

Introduction

Paragonimiasis is a neglected tropical disease (NTD) caused by the lung fluke Paragonimus westermani. It is transmitted by the ingestion of raw or inadequately cooked crabs. It is estimated that 21 million people have paragonimiasis, and 293 million people are at risk for infection [1]. In the Philippines, paragonimiasis is known to be endemic in at least 12 provinces [2]. There is a lack of a global program and guidelines specific to paragonimiasis control. While national policies concerning paragonimiasis control exist in the Philippines [3,4], these may need updating and further enhancement. For instance, paragonimiasis control in the country is tucked in a broad policy on Food and Waterborne Diseases Prevention and Control Program [3], which also includes diseases such as amebiasis among others.

Misdiagnosis of paragonimiasis as pulmonary tuberculosis (TB) and lack of mapping of endemic areas are continuing challenges in paragonimiasis surveillance and control. Diagnosis of paragonimiasis relies on the demonstration of Paragonimus ova using microscopy because immunological and molecular techniques are still at the experimental stage [5]. Diagnosis is complicated by similar clinical manifestations of paragonimiasis with TB. Chronic cough and hemoptysis, for example, are signs of both diseases. Chest radiographs may be unable to distinguish paragonimiasis from TB [6,7]. Misdiagnosis of paragonimiasis as non-response to TB treatment or even multi-drug resistant TB is not uncommon, especially in co-endemic areas [5,8,9]. In Sorsogon Province in the Philippines, for instance, up to 25% of the 252 patients in the rural health unit clinically diagnosed to have TB and who were unresponsive to TB treatment were subsequently diagnosed as cases of paragonimiasis [10]. Epidemiological data on paragonimiasis also remain limited and mapping of endemic areas has not been completed in the country. For instance, paragonimiasis cases have been reported in areas not declared as endemic, such as Zamboanga Sibugay [9].

Several studies on paragonimiasis surveillance and control have been conducted recently. A study in Lao People’s Democratic Republic showed the utility of a questionnaire in identifying paragonimiasis-endemic foci [11]. The Ziehl–Neelsen Staining technique (ZNS), which is used in TB diagnosis, was found to be capable of detecting Paragonimus ova [12]. A model for integrating paragonimiasis surveillance and control with the National Tuberculosis Control Program, on the other hand, has been developed, tested, and implemented in selected areas of the Philippines [9,13]

Despite these recent studies, continuing challenges in paragonimiasis surveillance and control remain. The utility of the aforementioned questionnaire in identifying paragonimiasis-endemic foci has not been determined in the Philippines. There is likewise a need to determine if local health workers, particularly medical technologists, can use the ZNS to detect Paragonimus ova. Testing of a questionnaire and the ZNS for the rapid identification of paragonimiasis-endemic foci may be used in updating policies and improving practice on paragonimiasis surveillance and control. It may likewise help in updating epidemiological data on paragonimiasis.

This study aimed to (1) develop a questionnaire to identify suspected paragonimiasis-endemic foci, (2) describe the epidemiology of paragonimiasis, and (3) evaluate ZNS in detecting Paragonimus ova.

Materials and methods

Development of the questionnaire

A series of workshops were conducted by the project team to develop the questionnaire for the rapid identification of suspected paragonimiasis-endemic foci. Criteria for identifying barangays (villages) which are suspected paragonimiasis-endemic foci [9] were considered in developing the questionnaire. The questionnaire was finalized in consultation with the Department of Health Zamboanga Peninsula Center for Health Development (DOH ZP CHD) and selected local government units (LGUs) in Zamboanga Region.

The tool, which was filled out by the municipal health officers, included five criteria for identifying barangays which are suspected paragonimiasis-endemic foci [9]. Suspected paragonimiasis-endemic foci were defined as areas that satisfied the inclusion criteria but indigenous cases are not yet confirmed by an active case finding. The inclusion criteria were (1) proximity of the barangay to a body of freshwater, (2) accessibility, peace and order situation, and cooperation of LGU, (3) consumption of raw or insufficiently cooked crustaceans by residents in the community, (4) reports of residents with productive cough with or without hemoptysis for at least 2 weeks or TB patients not responding to treatment or with recurrence of symptoms, and (5) confirmed cases of paragonimiasis in the last three years on records review. The questionnaire was used to identify the study sites or suspected paragonimiasis-endemic foci.

Study design, sites, and population

The study utilized a cross-sectional design which involved an examination of patients for TB and paragonimiasis using the integrated TB-paragonimiasis surveillance and control model [9] through active case finding in the barangays. This was conducted in suspected endemic areas in three provinces of Zamboanga Region, Philippines, namely Zamboanga del Norte, Zamboanga del Sur, and Zamboanga Sibugay. The region was selected due to previous paragonimiasis cases found in barangays in selected municipalities, as well as the strong partnership established between the DOH ZP CHD, University of the Philippines (UP) Manila, and various LGUs in the region. The questionnaire developed for the identification of suspected paragonimiasis-endemic foci was sent to rural health units, filled out by municipal health officers, and returned to the UP Manila Project Team through the DOH ZP CHD and respective provincial health offices. Barangays suspected as paragonimiasis-endemic foci were identified by the UP Manila Project Team using the responses on the questionnaires. Nine barangays from four different municipalities were selected as sites for active case finding (Table 1).

Table 1.

List of selected areas for active case finding of TB and paragonimiasis.

Province	Municipality	Barangay
Zamboanga Del Norte	Kalawit	Botong
		Daniel Maing
	Tampilisan	Balacbaan
		Banbanan
		Situbo
Zamboanga Del Sur	Tambulig	Bagong Kauswagan
		Dimalinao
		Libato
Zamboanga Sibugay	Tungawan	Little Margos

Sample size estimation

A total of 384 participants (96 per municipality) were required to estimate the prevalence of paragonimiasis given that the prevalence is within (6.7 ± 5%) in this population [13] with 95% confidence level.

To evaluate the sensitivity and specificity of ZNS in detecting Paragonimus ova given that the sensitivity of ZNS is 76.9% [12], the margin of error at 12%, and the prevalence of paragonimiasis is 6.7%, a total of 707 participants were required to be included in the study.

Model for integrating capacity building, surveillance, and research

A model for the integrated capacity building of health workers with surveillance and research of parasitic NTDs was implemented in this project. Medical technologists underwent a five-day training on diagnostic parasitology, which was integrated with active surveillance in the community. The training for medical technologists included pre and posttests, didactics, laboratory sessions, and practicum/fieldwork.

The first 2 days of the training on diagnostic parasitology for medical technologists included pretests, didactics, and laboratory sessions. Pretests, which consisted of a theoretical examination, where participants answered questions related to parasitic NTDs, and a practical test, where participants identified the parasites in the electronic images being shown, were administered to assess the participants’ knowledge on diagnosis, prevention, and control of parasitic NTDs. Lectures on epidemiology and control, laboratory diagnosis, and quality assurance in laboratory diagnosis were provided. The laboratory session, on the other hand, included practice and graded stool and sputum unknown exercises where participants examined preserved stool and sputum specimens for parasite ova.

The third and fourth days consisted of a fieldwork which served as the practicum of the participants. The fieldwork was supervised by faculty and trained research staff from UP Manila. Demonstration and return-demonstration of laboratory procedures were conducted. The participants also joined in the active surveillance. Symptomatic patients suspected to have paragonimiasis were recruited during the active surveillance. The specimens were collected, processed and, examined using the ZNS and NaOH concentration technique [14] for sputum specimens by the trained participants.

The last day included posttest, course evaluation, and awarding of certificates. The results of the posttest were compared with the pretests. The participants who completed the training were given a Certificate of Training with corresponding Continuing Professional Development units. The participants with the most improved score, as well as the participants who garnered the highest scores, were recognized.

Active surveillance

The proposed model for integrated TB-paragonimiasis surveillance and control [9] was modified in this project to include the examination of AFB smears prepared using the ZNS for Paragonimus ova detection under low power objective.

The UP Manila Project Team, through DOH ZP CHD, coordinated with the provincial health offices and rural health units for the active case finding in the nine barangays. The trained medical technologists, together with UP Manila project staff and DOH personnel, conducted the active case finding in the barangays. Case record form of the patients was filled out by trained research staff. All patients in the selected barangays who satisfied the inclusion criteria (i.e. cough of at least 2 weeks in duration) were included in the physical and the laboratory examinations. Prerequisites for their participation included submission of signed informed consent forms and two sputum samples (an early morning and a spot sputum sample) on the day of the consultation. Patient identifiers such as name, birth date, age, and sex of the participants were recorded by the assigned research staff. The specimens were processed and examined using the ZNS for detection of AFB and Paragonimus ova, while Sodium Hydroxide (NaOH) concentration technique was used for the detection of Paragonimus ova alone. The trained medical technologists examined the processed slides in the field laboratory set up in the barangay or in the Provincial DOH Office.

Quality assurance

The accuracy and reliability of sputum examination were ensured through quality assurance measures. This involved the proper collection of specimens, use of freshly prepared reagents, appropriate laboratory techniques, meticulous examination of processed specimens, and accurate reporting of findings. All slides positive for Paragonimus ova were validated by a reference or senior microscopist from UP Manila College of Public Health (CPH) Department of Parasitology (DOP). Electronic images of Paragonimus ova under the microscope were sent for verification by experts from the UP Manila CPH DOP. Quality control of AFB microscopy was performed by UP Manila CPH Department of Medical Microbiology. AFB slides included in the quality control for AFB were reexamined for Paragonimus ova by a reference microscopist from UP Manila CPH DOP who was blinded to the initial results.

Data processing and analysis

Collected data from the accomplished case record forms were encoded using Epi Info 7. Verification of data entries with source documents was done to ensure accuracy. Double encoding and data comparison were performed using the Data Compare module of Epi Info version 3.5.4. Data processing and statistical analysis were performed using STATA 12 (STATA 12®, StataCorp, Texas, USA). Descriptive statistics, which included the prevalence of paragonimiasis and TB, were calculated. Prevalence of TB among the patients was determined using ZNS technique. On the other hand, the prevalence of paragonimiasis was determined using both ZNS and NaOH concentration techniques. Sensitivity and specificity of the ZNS in detecting Paragonimus ova were calculated using the NaOH concentration technique as the reference standard. A 95% confidence interval was computed for prevalence estimates.

Ethical considerations

The study protocol was submitted and approved by the UP Manila – Research Ethics Board (UPMREB 2009-023-01) and the World Health Organization – Western Pacific Regional Office Ethics Review Committee (WHO ID: 2016.35.PHL.5.MVP). The study conformed with the national and international ethical guidelines to protect human subjects. Confidentiality was maintained through the replacement of participant identifiers with codes. Only the research team had access to the results to guarantee the anonymity of study participants and the confidentiality of information gathered. All study participants found positive for paragonimiasis and/or TB were referred to the rural health units for appropriate management following DOH guidelines. Feedback of the results of the study and recommendations were provided by UP Manila Project Team to the concerned provincial health offices and rural health units through the DOH ZP CHD at the end of the study.

Results

Utility of the questionnaire

Nine barangays satisfied the inclusion criteria in the questionnaire, of which five barangays are in Zamboanga del Norte, three in Zamboanga del Sur, and one in Zamboanga Sibugay. Five of the barangays have been included in the previous active case finding of the UP Manila Project Team. Paragonimiasis cases were found in seven out of the nine barangays which satisfied the inclusion criteria.

Epidemiology of paragonimiasis and TB

Out of the 373 patients examined, three (0.80%) were positive for TB, while 29 (7.77%) were positive for paragonimiasis. TB cases were found in Zamboanga del Norte, particularly in Daniel Maing in Kalawit (one case) and in Banbanan in Tampilisan (two cases). On the other hand, Libato in Tambulig, Zamboanga del Sur had the highest paragonimiasis prevalence at 26.67% (12 out of 45), while no case was found in the barangays of Banbanan in Tampilisan, Zamboanga del Norte and in Little Margos in Tungawan, Zamboanga Sibugay. Among the municipalities, Tambulig, Zamboanga del Sur had the highest paragonimiasis prevalence at 22.06%, while no case was found in Tungawan, Zamboanga Sibugay. Out of the 29 who were positive for paragonimiasis, all were positive using the NaOH concentration technique, while only eight were positive for paragonimiasis using ZNS (Table 2).

Table 2.

Prevalence of tuberculosis and paragonimiasis and positivity for Paragonimus ova using ZNS and NaOH concentration technique in selected barangays in Zamboanga Region (July 2017).

		Prevalence		Positive for Paragonimus
Province/Municipality/Barangays	Number of patients examined	TB (ZNS) No. (%)	Paragonimiasis (ZNS or NaOH) No. (%)	ZNS No. (%)	NaOH Conc. Technique No. (%)
Zamboanga Del Norte
Kalawit
Botong	63	0 (0.00)	5 (7.94)	2 (3.17)	5 (7.94)
Daniel Maing	51	1 (1.96)	2 (3.92)	2 (3.92)	2 (3.92)
Subtotal (Kalawit)	114	1 (0.88)	7 (6.14)	4 (3.51)	7 (6.14)
95% CI of prevalence		(0.05–5.50)	(2.93–12.39)	(1.13–9.27)	(2.72–12.69)
Tampilisan
Balacbaan	16	0 (0.00)	3 (18.75)	0 (0.00)	3 (18.75)
Banbanan	72	2(2.78)	0 (0.00)	0 (0.00)	0 (0.00)
Situbo	65	0 (0.00)	4 (6.15)	0 (0.00)	4 (6.15)
Subtotal (Tampilisan)	153	2 (1.31)	7 (4.58)	0 (0.00)	7 (4.58)
95% CI of prevalence		(0.23–5.13)	(2.19–9.33)		(2.02–9.56)
Subtotal (Zamboanga del Norte)	267	3 (1.12)	14 (5.24)	0 (0.00)	14 (5.24)
95% CI of prevalence		(0.29–3.52)	(3.12–8.68)		(3.01–8.83)
ZAMBOANGA DEL SUR
Tambulig
Bagong Kauswagan	17	0 (0.00)	2 (11.76)	0 (0.00)	2 (11.76)
Dimalinao	6	0 (0.00)	1 (16.67)	0 (0.00)	1 (16.67)
Libato	45	0 (0.00)	12 (26.67)	4 (8.89)	12 (26.67)
Subtotal (Tambulig, Zamboanga del Sur)	68	0 (0.00)	15 (22.06)	4 (5.88)	15 (22.06)
95% CI of prevalence			(13.69–33.56)	(1.90–15.14)	(13.26–34.06)
ZAMBOANGA SIBUGAY
Tungawan- Little Margos	38	0 (0.00)	0 (0.00)	0 (0.00)	0 (0.00)
Overall	373	3 (0.80) (0.21–2.53)	29 (7.77) (5.44–10.98)	8 (2.14) (1.00–4.35)	29 (7.77) (5.36–11.10)

The age group with the highest proportion of paragonimiasis-positive patients was 0–15 years old at 16.67% (3 out of 18), while it was lowest in the >60 years old age group at 4.62% (3 out of 65). It must be noted, however, that 124 (33.24%) of the patients had no information on their age, of which 4.03% were positive (5 out of 124) (Table 3). The prevalence of paragonimiasis was higher among males at 12.10% (15 out of 124), while 8.23% of females (13 out of 158) had paragonimiasis. It must also be noted that 91 (24.40%) of the patients had no record for sex, of which one is paragonimiasis positive (Table 3).

Table 3.

Age and sex distribution of patients in Zamboanga Region (July 2017).

Categories	No. of patients examined	Positive for TB (ZNS) No. (%)	Positive for Paragonimus ova (ZNS or NaOH) No. (%)
Age group
0–15	18	0	3 (16.67)
16–30	44	1 (2.27)	5 (11.36)
31–45	57	0	6 (10.53)
46–60	65	1 (1.54)	7 (10.77)
>60	65	0	3 (4.62)
No data	124	1 (0.81)	5 (4.03)
Sex
Male	124	0 (0.0)	15 (12.10)
Female	158	2 (1.27)	13 (8.23)
No data	91	1 (1.10)	1 (1.10)

Evaluation of ZNS in detecting Paragonimus ova

Using NaOH concentration technique as the reference standard for detecting Paragonimus ova, ZNS correctly detected 8 out of the 29 samples positive for Paragonimus ova, resulting in a sensitivity of 27.59% (95% CI: 12.73–47.24%). On the other hand, it correctly detected the 334 samples negative for Paragonimus ova, resulting in a specificity of 100% (95% CI: 98.90% to 100%). Figure 1 shows a Paragonimus egg as seen from a smear prepared using NaOH concentration technique, while Figure 2 and 3 shows Paragonimus eggs as seen from the ZNS.

Figure 1.

Paragonimus ovum in NaOH under HPO.

Figure 2.

Paragonimus ova in ZNS under HPO.

Figure 3.

Paragonimus ovum in ZNS under HPO.

Comparison of early morning and spot sputum samples in detecting AFB and Paragonimus ova

Two cases of paragonimiasis were only diagnosed using early morning samples, while three were only diagnosed using spot samples. On the other hand, one case of TB was diagnosed only by examining spot sputum samples while another was diagnosed by examining early morning samples.

Discussion

Utility of the questionnaire in identifying suspected paragonimiasis-endemic foci

Paragonimiasis cases were found in the barangays identified by the questionnaire as suspected paragonimiasis-endemic foci, except for Banbanan and Little Margos. This supports the finding of a study that a questionnaire is useful in identifying paragonimiasis-endemic foci [11]. The questionnaire, however, may be further improved. The criteria may be vague, and there is no documentation required on the part of the municipal health officers to support the responses. There may be a need to exclude ‘accessibility, peace and order situation, and cooperation of LGU’ in the criteria because an area will be endemic regardless of accessibility, peace and order situation, and LGU cooperation. Lastly, the questionnaire was developed based mostly on expert opinion. Systematic reviews, expert panel, and pilot in several endemic and non-endemic barangays may be used in questionnaire enhancement and further development.

Epidemiology of paragonimiasis

Similar to the study in the same region in 2015 [9], paragonimiasis cases were found in barangays not known to be endemic, such as Balacbaan in Tampilisan, Zamboanga del Norte and in Bagong Kauswagan, Dimalinao, and Libato in Tambulig, Zamboanga del Sur. Failure to classify an endemic area as such will deprive the population at risk, especially in hard-to-reach areas, the access to proper diagnosis and potentially life-saving treatment they need. It may likewise facilitate continuing transmission. This highlights the need to map the endemic areas and conduct periodic surveillance.

Compared to earlier findings in 2015, more paragonimiasis cases were found in the barangays of Botong in Kalawit and Situbo in Tampilisan, while one fewer case was found in Daniel Maing in Kalawit and Little Margos in Tungawan. Whether or not these differences were significant, however, was not determined. The prevalence of paragonimiasis is also considerably higher than the prevalence of TB in all study sites.

Areas such as Libato in Tambulig, which had the highest paragonimiasis prevalence, may require other prevention and control strategies, such as mass drug administration (MDA). WHO recommends MDA of triclabendazole (20 mg/kg) for areas that are highly endemic for paragonimiasis (reference). The challenge, however, is that no threshold for highly endemic areas and MDA guidelines, in general, have been established [5]. Moreover, the DOH still uses praziquantel for the treatment of paragonimiasis, as of project implementation. MDA of triclabendazole has been shown to be safe and effective in controlling fascioliasis [15]. The experience in MDA of triclabendazole for fascioliasis control may be a guide in assessing the effect of MDA of triclabendazole for paragonimiasis control.

The age and sex distribution of the paragonimiasis-positive patients suggest the need to reexamine the risk factors. Past studies suggested that paragonimiasis may occur mostly in adult males, possibly due to their drinking habits where they often eat raw crabs alongside the alcoholic drinks [16]. This study, however, showed that children may also be at risk for paragonimiasis and that the disease is almost as common in females as in males. This also supports the findings in an earlier study [10]. Obtaining reliable risk estimates of the risk factors may allow more targeted surveillance and control interventions.

Evaluation of ZNS in detecting Paragonimus ova

The ability of ZNS to detect Paragonimus ova will be useful to TB and paragonimiasis surveillance and control. It will decrease the chances of paragonimiasis being misdiagnosed as TB, which unnecessarily consumes TB control resources with no improvement in health outcomes. It will also allow the finding and treatment of more paragonimiasis cases, thereby reducing morbidity and transmission. ZNS also inactivates pathogens and may allow post-surveillance quality control of the specimens [9,12], both of which may not be possible with the NaOH concentration technique. While Gene Xpert may supersede ZNS in TB surveillance considering the former’s higher sensitivity and specificity and its ability to detect MDR TB [17], a roll-out of Gene Xpert in the health centers and villages may be unlikely in the near future especially in remote areas where paragonimiasis is often endemic. This is due to challenges with program funding, complete solution package, and the health system [18]. Thus, ZNS will still be used in surveillance in the coming years.

More accurate estimates of sensitivity and specificity of ZNS and other paragonimiasis diagnostics may be obtained by using complement fixation, considered by CDC as a standard diagnostic test for paragonimiasis [19], as a reference standard. Complement fixation and other serological and molecular techniques, however, are still considered experimental [5] and are thus not universally accepted as a reference standard for diagnosis of paragonimiasis. In the absence of a reference standard, sensitivity, and specificity of diagnostic techniques may be estimated using different statistical modeling techniques such as latent class analysis, a statistical method for identifying unmeasured class membership among subjects using categorical and/or continuous observed variables [20]. The utility of latent class analysis in estimating the sensitivity and specificity of a diagnostic technique in the absence of a reference standard has been demonstrated in several studies [21,22].

ZNS detected eight paragonimiasis cases, which is less than a third of the 29 paragonimiasis cases detected using the NaOH concentration technique. The results are consistent with the findings of a study where ZNS had a lower sensitivity than the concentration techniques [12]. If NaOH concentration technique is assumed as the reference standard, the sensitivity of ZNS will be lower than that described by an earlier study [12]. The lower detection rate may be because the ZNS is not a concentration technique. The ZNS may be modified to include concentration to increase the probability of detecting Paragonimus ova, especially in co-endemic areas. One modification of ZNS which can be explored is the bleach concentration technique [23]. While this technique was found to alter the morphology of Paragonimus ova, the ova remained identifiable [12]. The NaOH concentration technique and possibly other methods with equal, if not better, sensitivity and specificity for detecting Paragonimus ova may still be used until the ZNS is proven superior to these techniques.

Early morning and spot samples have varying results, emphasizing the importance of using both samples in surveillance. Modifying the procedures for ZNS and NaOH concentration technique, for instance, pooling and concentrating the early morning and spot samples prior to examination, may also be considered.

Feasibility of integrating capacity building, surveillance, and research

The study demonstrated the feasibility of integrating the capacity building with surveillance, and research. This innovative approach offers several advantages. First, it may reduce the cost of capacity building, surveillance, and research since the funds needed for these activities may be pooled. Second, this may help ensure that local health workers are capacitated to diagnose NTDs such as paragonimiasis. Third, it may help provide up-to-date data which may otherwise not be obtained due to the lack of surveillance. Lastly, it encourages the involvement of frontline LGUs and other stakeholders which is crucial toward effective disease control.

Proposed next steps for paragonimiasis surveillance and control

There is a need to adopt more sensitive and cost-effective diagnostics considering that microscopy of sputum samples is known to have poor sensitivity. Such diagnostics may include molecular techniques such as polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP). While further research may still be needed before more sensitive and cost-effective diagnostic techniques are used by the national program for paragonimiasis diagnosis and surveillance, ensuring the quality of available diagnostic techniques may be needed. Ensuring the availability of the needed equipment and supplies, as well as continuing capacity building for medical technologists through the collaboration of academe, DOH, and LGUs, are thus crucial.

Mapping of endemic areas and the implementation of a surveillance system are necessary for paragonimiasis control. Integrating paragonimiasis surveillance and control with the National Tuberculosis Control Program, rather than with the Food and Waterborne Diseases Prevention and Control Program, may be more advantageous since TB and paragonimiasis surveillance use the same sample, equipment (i.e. microscope), and possibly the same technique considering that ZNS can detect Paragonimus ova. In addition, the diagnosis of TB and paragonimiasis could be done by the health worker in the same point-of-care. This will thus maximize the use of resources and lessen the strain on the local health system. The model for integrated TB-paragonimiasis surveillance and control provides evidence on how paragonimiasis surveillance and control may be integrated with the National Tuberculosis Control Program [9].

Limitations

The barangays which did not satisfy the inclusion criteria were not included in the survey limits the study in evaluating the questionnaire. In theory, the study can calculate the positive predictive value of the questionnaire in identifying suspected paragonimiasis-endemic foci. The low number of barangays included, however, may make the positive predictive value estimate inaccurate.

The sample of the population obtained may not be random because only symptomatic patients who presented to the field laboratory were included. The target sample size, likewise, was not met. These may have limited the external validity of the study.

Some patients had only submitted early morning samples especially for patients who live far from the field laboratory and had to send their samples through the health workers. There were also patients who had only submitted spot sputum samples, particularly those who were unable to collect their early morning sputum samples when they woke up. Likewise, there were samples that were only processed and examined using ZNS especially those samples with insufficient amount. Lastly, there were samples that were only processed using NaOH concentration technique especially those which were salivary in consistency since the medical technologists are unable to examine these ZNS slides. These resulted in the different number of spot and sputum samples, as well as a different number of the samples processed using ZNS and/or NaOH concentration technique.

Some patients had incomplete information on age and sex due to factors such as the inherent challenges encountered in the conduct of the fieldwork. For example, some of the samples were submitted by the patients through the health workers (instead of the patients appearing personally in the field laboratory) due to the inaccessibility of the barangays. This made it difficult to generalize on the age and sex distribution of those who were positive for TB and paragonimiasis.

The lack of accepted reference standard for diagnosis of paragonimiasis meant that the computed sensitivity and specificity may not be accurate. There is also a possibility of confirmation bias, i.e. a microscopist may search for Paragonimus ova more meticulously in ZNS if a patient is positive for Paragonimus ova using the NaOH concentration technique.

Further studies

Future research areas identified by the project include the optimization of the questionnaire for identifying paragonimiasis-endemic foci, evaluating ZNS for detecting Paragonimus ova using statistical modeling techniques, and evaluation of modified ZNS techniques, such as bleach concentration technique for detecting Paragonimus ova. The cost-effectiveness of an integrated TB-paragonimiasis surveillance and control and the feasibility of MDA of triclabendazole in highly endemic areas for paragonimiasis could be studied to guide policy formulation and program implementation. Mapping of paragonimiasis-endemic areas and obtaining reliable estimates of the risk factors will also support paragonimiasis control.

Conclusion

The simple questionnaire for identifying suspected paragonimiasis-endemic foci was useful in identifying endemic barangays. Further improvements on the questionnaire may include refining the inclusion criteria, e.g. removing ‘accessibility, peace and order situation, and cooperation of LGU’, and requiring documentation to support the responses on the questionnaire. Mapping and surveillance of the endemic barangays are needed to guide efforts toward disease prevention and control. Risk factors for paragonimiasis may be further studied. The ZNS can detect Paragonimus ova although it has low sensitivity. Modifying the procedures of ZNS, for instance concentrating the sputum sample, may be considered to improve the detection of Paragonimus ova. Using ZNS to detect Paragonimus ova is advantageous because it is already used in the National Tuberculosis Control Program, inactivates pathogens, and may allow post-surveillance quality control, all of which may not be possible with the NaOH concentration technique. It also supports the integration of surveillance and control of TB and paragonimiasis. Until the sensitivity of ZNS is improved, the NaOH concentration technique and other methods with equal, if not higher sensitivity, may still be used.

Funding Statement

The research was funded through the Joint TDR/World Health Organization Western Pacific Region Small Grants Scheme 2016 (Project No. HQTDR1611221).

Acknowledgments

The authors would like to extend their sincere gratitude to DOH ZP CHD; provincial LGUs of Zamboanga del Norte, Zamboanga del Sur, and Zamboanga Sibugay; and municipal LGUs of Kalawit, Tampilisan, Tambulig, and Tungawan, as well as to Ms. Jessa Flores, for their valuable support in the implementation of the study. The authors would also like to thank the Special Programme for Research and Training in Tropical Diseases (TDR) and the World Health Organization Western Pacific Region for providing financial support to implement the study.

Disclosure statement

The researchers declare no conflict of interest.