The Human Filaria Loa loa: Update on Diagnostics and Immune Response

Abstract

Loa loa loiasis was considered an anecdotal disease 30 years ago. Its spread in Equatorial Africa and the side effects associated with mass drug administration programs against filariasis in co-endemic areas have drawn the attention of the international research community. Progress in research conducted to date has provided insight into the immunobiology of this parasite. An interesting finding reported in several studies is that 70% of individuals with loiasis do not carry microfilariae in their blood, and 30% are microfilaremic, suggesting the involvement of several immunological mechanisms, as shown by elevated specific IgG4 and IgE levels signifying a potential cross-linking mechanism between the two isotypes via L. loa antigen to prevent allergy. A mechanism of anergy in the appearance of microfilariae in the peripheral blood results in immunological unresponsiveness in individuals with microfilariae. There is an interaction between other pathogens (parasites, bacteria, viruses) in individuals co-infected with L. loa. The strong antigen cross-reactivity between L. loa and lymphatic filarial worms warrants a re-evaluation of the distribution of the latter in co-endemic regions. The mechanism of concomitant immunity observed in the elimination of microfilariae or infective larvae (third-stage larvae, L3) may be used for the conception of an immunoprophylactic strategy.

Keywords: Loa loa; cross-reactivity; diagnostics; immune anergy; immune evasion.

Figure 1

Loa loa microfilariae. Microfilariae purified after Percoll gradient cell separation (A) and May–Grünwald Giemsa staining (B and C). The sheath can be seen as an extended translucid structure along the end tail (arrows).

Figure 2

Chrysops vector of Loa loa. View of Chrysops silacea with the typical two bars on the back (A) and Chrysops dimidiata (B). Ventral side of the two flies, C. silacea (C) and C. dimidiate (D). These flies can carry more infective L3 along their proboscis and thorax. These serve as a source of natural infective-stage larvae of Loa loa.

Figure 3

Potential mechanism of blocking allergic reactions in loiasis. Cross-linkage between IgE/IgG4 (via L. loa antigens) and fixation on FcεRI (IgE) and FcγRII by IgG4, resulting in the absence of allergic reaction.

Figure 4

Potential mechanism of anergy in loiasis. Normal antigen presentation (A) involves three signals with antigen (Ag) recognition. In microfilaremic loiasis, the co-stimulation signal (B) with B7 associated with CTLA-4 in lymphocytes and IL-10 synthesis is not adequate.

Figure 5

Three-dimensional Loa loa microfilaria sheath protein generated with the Swiss-Prot bioinformatics program. The molecular weight is 17.225 kDa, with a PI of 5.80 containing a valine-rich region, leucine slipper, phosphorylation site, myristoylation site, and glycosylation site, with alternating hydrophobic and hydrophilic regions.

Figure 6

Indices of concomitant immunity. Microfilariae and specific IgG4 prevalence plotted against age. As shown, both increase up to a certain age, after which microfilariae start decreasing while specific IgG4 remains high, suggesting a continuous stimulation either by adult resident worms or new infective L3 without establishment of a new infection.

Figure 7

Isolation and purification of microfilariae by Percoll gradients. Iso-osmotic Percoll was prepared by adding 9 parts (v/v) Percoll to RPMI-1640 MEDIUM 10x (SIP), and adding 2 mL gradient solution of 40%, 50%, 65% SIP in a 15-mL polystyrene tube, respectively, over the layer of these gradient solutions. Subsequently, 2 mL whole blood from an infected individual was added and centrifuged at 1000 g for 20 min. Layers 3 and 4 containing microfilariae were isolated and passed through a 5-µm filter. The motile microfilariae were then released in RPMI-1640 MEDIUM 1x (Van Hoegaerden and Ivanoff, 1986). These microfilariae are a source of crude antigens and genomic DNA.

Figure 8

Infective larvae L3. Infective larvae isolated via the Baerman technique in naturally infected Chrysops. Some flies can harbor one (A) or more than 100 larvae (B) of L. loa L3, with a mean length of 18 mm, seen with the naked eye or under binocular loupe.

Figure 9

Baerman technique and device. A funnel with a stopcock at the end is filled with HBSS 1x and small bag of crushed Chrysops captured in different villages. After 10 min of incubation in the HBSS solution, the stopcock is opened and HBSS containing L3 is harvested in a Petri dish under the funnel.